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Raffi Hovasapian

Raffi Hovasapian

Amino Acid Sequencing of a Peptide Chain

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Table of Contents

Section 1: Preliminaries on Aqueous Chemistry
Aqueous Solutions & Concentration

39m 57s

Intro
0:00
Aqueous Solutions and Concentration
0:46
Definition of Solution
1:28
Example: Sugar Dissolved in Water
2:19
Example: Salt Dissolved in Water
3:04
A Solute Does Not Have to Be a Solid
3:37
A Solvent Does Not Have to Be a Liquid
5:02
Covalent Compounds
6:55
Ionic Compounds
7:39
Example: Table Sugar
9:12
Example: MgCl₂
10:40
Expressing Concentration: Molarity
13:42
Example 1
14:47
Example 1: Question
14:50
Example 1: Solution
15:40
Another Way to Express Concentration
22:01
Example 2
24:00
Example 2: Question
24:01
Example 2: Solution
24:49
Some Other Ways of Expressing Concentration
27:52
Example 3
29:30
Example 3: Question
29:31
Example 3: Solution
31:02
Dilution & Osmotic Pressure

38m 53s

Intro
0:00
Dilution
0:45
Definition of Dilution
0:46
Example 1: Question
2:08
Example 1: Basic Dilution Equation
4:20
Example 1: Solution
5:31
Example 2: Alternative Approach
12:05
Osmotic Pressure
14:34
Colligative Properties
15:02
Recall: Covalent Compounds and Soluble Ionic Compounds
17:24
Properties of Pure Water
19:42
Addition of a Solute
21:56
Osmotic Pressure: Conceptual Example
24:00
Equation for Osmotic Pressure
29:30
Example of 'i'
31:38
Example 3
32:50
More on Osmosis

29m 1s

Intro
0:00
More on Osmosis
1:25
Osmotic Pressure
1:26
Example 1: Molar Mass of Protein
5:25
Definition, Equation, and Unit of Osmolarity
13:13
Example 2: Osmolarity
15:19
Isotonic, Hypertonic, and Hypotonic
20:20
Example 3
22:20
More on Isotonic, Hypertonic, and Hypotonic
26:14
Osmosis vs. Osmotic Pressure
27:56
Acids & Bases

39m 11s

Intro
0:00
Acids and Bases
1:16
Let's Begin With H₂O
1:17
P-Scale
4:22
Example 1
6:39
pH
9:43
Strong Acids
11:10
Strong Bases
13:52
Weak Acids & Bases Overview
14:32
Weak Acids
15:49
Example 2: Phosphoric Acid
19:30
Weak Bases
24:50
Weak Base Produces Hydroxide Indirectly
25:41
Example 3: Pyridine
29:07
Acid Form and Base Form
32:02
Acid Reaction
35:50
Base Reaction
36:27
Ka, Kb, and Kw
37:14
Titrations and Buffers

41m 33s

Intro
0:00
Titrations
0:27
Weak Acid
0:28
Rearranging the Ka Equation
1:45
Henderson-Hasselbalch Equation
3:52
Fundamental Reaction of Acids and Bases
5:36
The Idea Behind a Titration
6:27
Let's Look at an Acetic Acid Solution
8:44
Titration Curve
17:00
Acetate
23:57
Buffers
26:57
Introduction to Buffers
26:58
What is a Buffer?
29:40
Titration Curve & Buffer Region
31:44
How a Buffer Works: Adding OH⁻
34:44
How a Buffer Works: Adding H⁺
35:58
Phosphate Buffer System
38:02
Example Problems with Acids, Bases & Buffers

44m 19s

Intro
0:00
Example 1
1:21
Example 1: Properties of Glycine
1:22
Example 1: Part A
3:40
Example 1: Part B
4:40
Example 2
9:02
Example 2: Question
9:03
Example 2: Total Phosphate Concentration
12:23
Example 2: Final Solution
17:10
Example 3
19:34
Example 3: Question
19:35
Example 3: pH Before
22:18
Example 3: pH After
24:24
Example 3: New pH
27:54
Example 4
30:00
Example 4: Question
30:01
Example 4: Equilibria
32:52
Example 4: 1st Reaction
38:04
Example 4: 2nd Reaction
39:53
Example 4: Final Solution
41:33
Hydrolysis & Condensation Reactions

18m 45s

Intro
0:00
Hydrolysis and Condensation Reactions
0:50
Hydrolysis
0:51
Condensation
2:42
Example 1: Hydrolysis of Ethyl Acetate
4:52
Example 2: Condensation of Acetic Acid with Ethanol
8:42
Example 3
11:18
Example 4: Formation & Hydrolysis of a Peptide Bond Between the Amino Acids Alanine & Serine
14:56
Section 2: Amino Acids & Proteins: Primary Structure
Amino Acids

38m 19s

Intro
0:00
Amino Acids
0:17
Proteins & Amino Acids
0:18
Difference Between Amino Acids
4:20
α-Carbon
7:08
Configuration in Biochemistry
10:43
L-Glyceraldehyde & Fischer Projection
12:32
D-Glyceraldehyde & Fischer Projection
15:31
Amino Acids in Biological Proteins are the L Enantiomer
16:50
L-Amino Acid
18:04
L-Amino Acids Correspond to S-Enantiomers in the RS System
20:10
Classification of Amino Acids
22:53
Amino Acids With Non-Polar R Groups
26:45
Glycine
27:00
Alanine
27:48
Valine
28:15
Leucine
28:58
Proline
31:08
Isoleucine
32:42
Methionine
33:43
Amino Acids With Aromatic R Groups
34:33
Phenylalanine
35:26
Tyrosine
36:02
Tryptophan
36:32
Amino Acids, Continued

27m 14s

Intro
0:00
Amino Acids With Positively Charged R Groups
0:16
Lysine
0:52
Arginine
1:55
Histidine
3:15
Amino Acids With Negatively Charged R Groups
6:28
Aspartate
6:58
Glutamate
8:11
Amino Acids With Uncharged, but Polar R Groups
8:50
Serine
8:51
Threonine
10:21
Cysteine
11:06
Asparagine
11:35
Glutamine
12:44
More on Amino Acids
14:18
Cysteine Dimerizes to Form Cystine
14:53
Tryptophan, Tyrosine, and Phenylalanine
19:07
Other Amino Acids
20:53
Other Amino Acids: Hydroxy Lysine
22:34
Other Amino Acids: r-Carboxy Glutamate
25:37
Acid/Base Behavior of Amino Acids

48m 28s

Intro
0:00
Acid/Base Behavior of Amino Acids
0:27
Acid/Base Behavior of Amino Acids
0:28
Let's Look at Alanine
1:57
Titration of Acidic Solution of Alanine with a Strong Base
2:51
Amphoteric Amino Acids
13:24
Zwitterion & Isoelectric Point
16:42
Some Amino Acids Have 3 Ionizable Groups
20:35
Example: Aspartate
24:44
Example: Tyrosine
28:50
Rule of Thumb
33:04
Basis for the Rule
35:59
Example: Describe the Degree of Protonation for Each Ionizable Group
38:46
Histidine is Special
44:58
Peptides & Proteins

45m 18s

Intro
0:00
Peptides and Proteins
0:15
Introduction to Peptides and Proteins
0:16
Formation of a Peptide Bond: The Bond Between 2 Amino Acids
1:44
Equilibrium
7:53
Example 1: Build the Following Tripeptide Ala-Tyr-Ile
9:48
Example 1: Shape Structure
15:43
Example 1: Line Structure
17:11
Peptides Bonds
20:08
Terms We'll Be Using Interchangeably
23:14
Biological Activity & Size of a Peptide
24:58
Multi-Subunit Proteins
30:08
Proteins and Prosthetic Groups
32:13
Carbonic Anhydrase
37:35
Primary, Secondary, Tertiary, and Quaternary Structure of Proteins
40:26
Amino Acid Sequencing of a Peptide Chain

42m 47s

Intro
0:00
Amino Acid Sequencing of a Peptide Chain
0:30
Amino Acid Sequence and Its Structure
0:31
Edman Degradation: Overview
2:57
Edman Degradation: Reaction - Part 1
4:58
Edman Degradation: Reaction - Part 2
10:28
Edman Degradation: Reaction - Part 3
13:51
Mechanism Step 1: PTC (Phenylthiocarbamyl) Formation
19:01
Mechanism Step 2: Ring Formation & Peptide Bond Cleavage
23:03
Example: Write Out the Edman Degradation for the Tripeptide Ala-Tyr-Ser
30:29
Step 1
30:30
Step 2
34:21
Step 3
36:56
Step 4
38:28
Step 5
39:24
Step 6
40:44
Sequencing Larger Peptides & Proteins

1h 2m 33s

Intro
0:00
Sequencing Larger Peptides and Proteins
0:28
Identifying the N-Terminal Amino Acids With the Reagent Fluorodinitrobenzene (FDNB)
0:29
Sequencing Longer Peptides & Proteins Overview
5:54
Breaking Peptide Bond: Proteases and Chemicals
8:16
Some Enzymes/Chemicals Used for Fragmentation: Trypsin
11:14
Some Enzymes/Chemicals Used for Fragmentation: Chymotrypsin
13:02
Some Enzymes/Chemicals Used for Fragmentation: Cyanogen Bromide
13:28
Some Enzymes/Chemicals Used for Fragmentation: Pepsin
13:44
Cleavage Location
14:04
Example: Chymotrypsin
16:44
Example: Pepsin
18:17
More on Sequencing Larger Peptides and Proteins
19:29
Breaking Disulfide Bonds: Performic Acid
26:08
Breaking Disulfide Bonds: Dithiothreitol Followed by Iodoacetate
31:04
Example: Sequencing Larger Peptides and Proteins
37:03
Part 1 - Breaking Disulfide Bonds, Hydrolysis and Separation
37:04
Part 2 - N-Terminal Identification
44:16
Part 3 - Sequencing Using Pepsin
46:43
Part 4 - Sequencing Using Cyanogen Bromide
52:02
Part 5 - Final Sequence
56:48
Peptide Synthesis (Merrifield Process)

49m 12s

Intro
0:00
Peptide Synthesis (Merrifield Process)
0:31
Introduction to Synthesizing Peptides
0:32
Merrifield Peptide Synthesis: General Scheme
3:03
So What Do We Do?
6:07
Synthesis of Protein in the Body Vs. The Merrifield Process
7:40
Example: Synthesis of Ala-Gly-Ser
9:21
Synthesis of Ala-Gly-Ser: Reactions Overview
11:41
Synthesis of Ala-Gly-Ser: Reaction 1
19:34
Synthesis of Ala-Gly-Ser: Reaction 2
24:34
Synthesis of Ala-Gly-Ser: Reaction 3
27:34
Synthesis of Ala-Gly-Ser: Reaction 4 & 4a
28:48
Synthesis of Ala-Gly-Ser: Reaction 5
33:38
Synthesis of Ala-Gly-Ser: Reaction 6
36:45
Synthesis of Ala-Gly-Ser: Reaction 7 & 7a
37:44
Synthesis of Ala-Gly-Ser: Reaction 8
39:47
Synthesis of Ala-Gly-Ser: Reaction 9 & 10
43:23
Chromatography: Eluent, Stationary Phase, and Eluate
45:55
More Examples with Amino Acids & Peptides

54m 31s

Intro
0:00
Example 1
0:22
Data
0:23
Part A: What is the pI of Serine & Draw the Correct Structure
2:11
Part B: How Many mL of NaOH Solution Have Been Added at This Point (pI)?
5:27
Part C: At What pH is the Average Charge on Serine
10:50
Part D: Draw the Titration Curve for This Situation
14:50
Part E: The 10 mL of NaOH Added to the Solution at the pI is How Many Equivalents?
17:35
Part F: Serine Buffer Solution
20:22
Example 2
23:04
Data
23:05
Part A: Calculate the Minimum Molar Mass of the Protein
25:12
Part B: How Many Tyr Residues in this Protein?
28:34
Example 3
30:08
Question
30:09
Solution
34:30
Example 4
48:46
Question
48:47
Solution
49:50
Section 3: Proteins: Secondary, Tertiary, and Quaternary Structure
Alpha Helix & Beta Conformation

50m 52s

Intro
0:00
Alpha Helix and Beta Conformation
0:28
Protein Structure Overview
0:29
Weak interactions Among the Amino Acid in the Peptide Chain
2:11
Two Principals of Folding Patterns
4:56
Peptide Bond
7:00
Peptide Bond: Resonance
9:46
Peptide Bond: φ Bond & ψ Bond
11:22
Secondary Structure
15:08
α-Helix Folding Pattern
17:28
Illustration 1: α-Helix Folding Pattern
19:22
Illustration 2: α-Helix Folding Pattern
21:39
β-Sheet
25:16
β-Conformation
26:04
Parallel & Anti-parallel
28:44
Parallel β-Conformation Arrangement of the Peptide Chain
30:12
Putting Together a Parallel Peptide Chain
35:16
Anti-Parallel β-Conformation Arrangement
37:42
Tertiary Structure
45:03
Quaternary Structure
45:52
Illustration 3: Myoglobin Tertiary Structure & Hemoglobin Quaternary Structure
47:13
Final Words on Alpha Helix and Beta Conformation
48:34
Section 4: Proteins: Function
Protein Function I: Ligand Binding & Myoglobin

51m 36s

Intro
0:00
Protein Function I: Ligand Binding & Myoglobin
0:30
Ligand
1:02
Binding Site
2:06
Proteins are Not Static or Fixed
3:36
Multi-Subunit Proteins
5:46
O₂ as a Ligand
7:21
Myoglobin, Protoporphyrin IX, Fe ²⁺, and O₂
12:54
Protoporphyrin Illustration
14:25
Myoglobin With a Heme Group Illustration
17:02
Fe²⁺ has 6 Coordination Sites & Binds O₂
18:10
Heme
19:44
Myoglobin Overview
22:40
Myoglobin and O₂ Interaction
23:34
Keq or Ka & The Measure of Protein's Affinity for Its Ligand
26:46
Defining α: Fraction of Binding Sites Occupied
32:52
Graph: α vs. [L]
37:33
For The Special Case of α = 0.5
39:01
Association Constant & Dissociation Constant
43:54
α & Kd
45:15
Myoglobin's Binding of O₂
48:20
Protein Function II: Hemoglobin

1h 3m 36s

Intro
0:00
Protein Function II: Hemoglobin
0:14
Hemoglobin Overview
0:15
Hemoglobin & Its 4 Subunits
1:22
α and β Interactions
5:18
Two Major Conformations of Hb: T State (Tense) & R State (Relaxed)
8:06
Transition From The T State to R State
12:03
Binding of Hemoglobins & O₂
14:02
Binding Curve
18:32
Hemoglobin in the Lung
27:28
Signoid Curve
30:13
Cooperative Binding
32:25
Hemoglobin is an Allosteric Protein
34:26
Homotropic Allostery
36:18
Describing Cooperative Binding Quantitatively
38:06
Deriving The Hill Equation
41:52
Graphing the Hill Equation
44:43
The Slope and Degree of Cooperation
46:25
The Hill Coefficient
49:48
Hill Coefficient = 1
51:08
Hill Coefficient < 1
55:55
Where the Graph Hits the x-axis
56:11
Graph for Hemoglobin
58:02
Protein Function III: More on Hemoglobin

1h 7m 16s

Intro
0:00
Protein Function III: More on Hemoglobin
0:11
Two Models for Cooperative Binding: MWC & Sequential Model
0:12
MWC Model
1:31
Hemoglobin Subunits
3:32
Sequential Model
8:00
Hemoglobin Transports H⁺ & CO₂
17:23
Binding Sites of H⁺ and CO₂
19:36
CO₂ is Converted to Bicarbonate
23:28
Production of H⁺ & CO₂ in Tissues
27:28
H⁺ & CO₂ Binding are Inversely Related to O₂ Binding
28:31
The H⁺ Bohr Effect: His¹⁴⁶ Residue on the β Subunits
33:31
Heterotropic Allosteric Regulation of O₂ Binding by 2,3-Biphosphoglycerate (2,3 BPG)
39:53
Binding Curve for 2,3 BPG
56:21
Section 5: Enzymes
Enzymes I

41m 38s

Intro
0:00
Enzymes I
0:38
Enzymes Overview
0:39
Cofactor
4:38
Holoenzyme
5:52
Apoenzyme
6:40
Riboflavin, FAD, Pyridoxine, Pyridoxal Phosphate Structures
7:28
Carbonic Anhydrase
8:45
Classification of Enzymes
9:55
Example: EC 1.1.1.1
13:04
Reaction of Oxidoreductases
16:23
Enzymes: Catalysts, Active Site, and Substrate
18:28
Illustration of Enzymes, Substrate, and Active Site
27:22
Catalysts & Activation Energies
29:57
Intermediates
36:00
Enzymes II

44m 2s

Intro
0:00
Enzymes II: Transitions State, Binding Energy, & Induced Fit
0:18
Enzymes 'Fitting' Well With The Transition State
0:20
Example Reaction: Breaking of a Stick
3:40
Another Energy Diagram
8:20
Binding Energy
9:48
Enzymes Specificity
11:03
Key Point: Optimal Interactions Between Substrate & Enzymes
15:15
Induced Fit
16:25
Illustrations: Induced Fit
20:58
Enzymes II: Catalytic Mechanisms
22:17
General Acid/Base Catalysis
23:56
Acid Form & Base Form of Amino Acid: Glu &Asp
25:26
Acid Form & Base Form of Amino Acid: Lys & Arg
26:30
Acid Form & Base Form of Amino Acid: Cys
26:51
Acid Form & Base Form of Amino Acid: His
27:30
Acid Form & Base Form of Amino Acid: Ser
28:16
Acid Form & Base Form of Amino Acid: Tyr
28:30
Example: Phosphohexose Isomerase
29:20
Covalent Catalysis
34:19
Example: Glyceraldehyde 3-Phosphate Dehydrogenase
35:34
Metal Ion Catalysis: Isocitrate Dehydrogenase
38:45
Function of Mn²⁺
42:15
Enzymes III: Kinetics

56m 40s

Intro
0:00
Enzymes III: Kinetics
1:40
Rate of an Enzyme-Catalyzed Reaction & Substrate Concentration
1:41
Graph: Substrate Concentration vs. Reaction Rate
10:43
Rate At Low and High Substrate Concentration
14:26
Michaelis & Menten Kinetics
20:16
More On Rate & Concentration of Substrate
22:46
Steady-State Assumption
26:02
Rate is Determined by How Fast ES Breaks Down to Product
31:36
Total Enzyme Concentration: [Et] = [E] + [ES]
35:35
Rate of ES Formation
36:44
Rate of ES Breakdown
38:40
Measuring Concentration of Enzyme-Substrate Complex
41:19
Measuring Initial & Maximum Velocity
43:43
Michaelis & Menten Equation
46:44
What Happens When V₀ = (1/2) Vmax?
49:12
When [S] << Km
53:32
When [S] >> Km
54:44
Enzymes IV: Lineweaver-Burk Plots

20m 37s

Intro
0:00
Enzymes IV: Lineweaver-Burk Plots
0:45
Deriving The Lineweaver-Burk Equation
0:46
Lineweaver-Burk Plots
3:55
Example 1: Carboxypeptidase A
8:00
More on Km, Vmax, and Enzyme-catalyzed Reaction
15:54
Enzymes V: Enzyme Inhibition

51m 37s

Intro
0:00
Enzymes V: Enzyme Inhibition Overview
0:42
Enzyme Inhibitors Overview
0:43
Classes of Inhibitors
2:32
Competitive Inhibition
3:08
Competitive Inhibition
3:09
Michaelis & Menten Equation in the Presence of a Competitive Inhibitor
7:40
Double-Reciprocal Version of the Michaelis & Menten Equation
14:48
Competitive Inhibition Graph
16:37
Uncompetitive Inhibition
19:23
Uncompetitive Inhibitor
19:24
Michaelis & Menten Equation for Uncompetitive Inhibition
22:10
The Lineweaver-Burk Equation for Uncompetitive Inhibition
26:04
Uncompetitive Inhibition Graph
27:42
Mixed Inhibition
30:30
Mixed Inhibitor
30:31
Double-Reciprocal Version of the Equation
33:34
The Lineweaver-Burk Plots for Mixed Inhibition
35:02
Summary of Reversible Inhibitor Behavior
38:00
Summary of Reversible Inhibitor Behavior
38:01
Note: Non-Competitive Inhibition
42:22
Irreversible Inhibition
45:15
Irreversible Inhibition
45:16
Penicillin & Transpeptidase Enzyme
46:50
Enzymes VI: Regulatory Enzymes

51m 23s

Intro
0:00
Enzymes VI: Regulatory Enzymes
0:45
Regulatory Enzymes Overview
0:46
Example: Glycolysis
2:27
Allosteric Regulatory Enzyme
9:19
Covalent Modification
13:08
Two Other Regulatory Processes
16:28
Allosteric Regulation
20:58
Feedback Inhibition
25:12
Feedback Inhibition Example: L-Threonine → L-Isoleucine
26:03
Covalent Modification
27:26
Covalent Modulators: -PO₃²⁻
29:30
Protein Kinases
31:59
Protein Phosphatases
32:47
Addition/Removal of -PO₃²⁻ and the Effect on Regulatory Enzyme
33:36
Phosphorylation Sites of a Regulatory Enzyme
38:38
Proteolytic Cleavage
41:48
Zymogens: Chymotrypsin & Trypsin
43:58
Enzymes That Use More Than One Regulatory Process: Bacterial Glutamine Synthetase
48:59
Why The Complexity?
50:27
Enzymes VII: Km & Kcat

54m 49s

Intro
0:00
Km
1:48
Recall the Michaelis–Menten Equation
1:49
Km & Enzyme's Affinity
6:18
Rate Forward, Rate Backward, and Equilibrium Constant
11:08
When an Enzyme's Affinity for Its Substrate is High
14:17
More on Km & Enzyme Affinity
17:29
The Measure of Km Under Michaelis–Menten kinetic
23:19
Kcat (First-order Rate Constant or Catalytic Rate Constant)
24:10
Kcat: Definition
24:11
Kcat & The Michaelis–Menten Postulate
25:18
Finding Vmax and [Et}
27:27
Units for Vmax and Kcat
28:26
Kcat: Turnover Number
28:55
Michaelis–Menten Equation
32:12
Km & Kcat
36:37
Second Order Rate Equation
36:38
(Kcat)/(Km): Overview
39:22
High (Kcat)/(Km)
40:20
Low (Kcat)/(Km)
43:16
Practical Big Picture
46:28
Upper Limit to (Kcat)/(Km)
48:56
More On Kcat and Km
49:26
Section 6: Carbohydrates
Monosaccharides

1h 17m 46s

Intro
0:00
Monosaccharides
1:49
Carbohydrates Overview
1:50
Three Major Classes of Carbohydrates
4:48
Definition of Monosaccharides
5:46
Examples of Monosaccharides: Aldoses
7:06
D-Glyceraldehyde
7:39
D-Erythrose
9:00
D-Ribose
10:10
D-Glucose
11:20
Observation: Aldehyde Group
11:54
Observation: Carbonyl 'C'
12:30
Observation: D & L Naming System
12:54
Examples of Monosaccharides: Ketose
16:54
Dihydroxy Acetone
17:28
D-Erythrulose
18:30
D-Ribulose
19:49
D-Fructose
21:10
D-Glucose Comparison
23:18
More information of Ketoses
24:50
Let's Look Closer at D-Glucoses
25:50
Let's Look At All the D-Hexose Stereoisomers
31:22
D-Allose
32:20
D-Altrose
33:01
D-Glucose
33:39
D-Gulose
35:00
D-Mannose
35:40
D-Idose
36:42
D-Galactose
37:14
D-Talose
37:42
Epimer
40:05
Definition of Epimer
40:06
Example of Epimer: D-Glucose, D-Mannose, and D-Galactose
40:57
Hemiacetal or Hemiketal
44:36
Hemiacetal/Hemiketal Overview
45:00
Ring Formation of the α and β Configurations of D-Glucose
50:52
Ring Formation of the α and β Configurations of Fructose
1:01:39
Haworth Projection
1:07:34
Pyranose & Furanose Overview
1:07:38
Haworth Projection: Pyranoses
1:09:30
Haworth Projection: Furanose
1:14:56
Hexose Derivatives & Reducing Sugars

37m 6s

Intro
0:00
Hexose Derivatives
0:15
Point of Clarification: Forming a Cyclic Sugar From a Linear Sugar
0:16
Let's Recall the α and β Anomers of Glucose
8:42
α-Glucose
10:54
Hexose Derivatives that Play Key Roles in Physiology Progression
17:38
β-Glucose
18:24
β-Glucosamine
18:48
N-Acetyl-β-Glucosamine
20:14
β-Glucose-6-Phosphate
22:22
D-Gluconate
24:10
Glucono-δ-Lactone
26:33
Reducing Sugars
29:50
Reducing Sugars Overview
29:51
Reducing Sugars Example: β-Galactose
32:36
Disaccharides

43m 32s

Intro
0:00
Disaccharides
0:15
Disaccharides Overview
0:19
Examples of Disaccharides & How to Name Them
2:49
Disaccharides Trehalose Overview
15:46
Disaccharides Trehalose: Flip
20:52
Disaccharides Trehalose: Spin
28:36
Example: Draw the Structure
33:12
Polysaccharides

39m 25s

Intro
0:00
Recap Example: Draw the Structure of Gal(α1↔β1)Man
0:38
Polysaccharides
9:46
Polysaccharides Overview
9:50
Homopolysaccharide
13:12
Heteropolysaccharide
13:47
Homopolysaccharide as Fuel Storage
16:23
Starch Has Two Types of Glucose Polymer: Amylose
17:10
Starch Has Two Types of Glucose Polymer: Amylopectin
18:04
Polysaccharides: Reducing End & Non-Reducing End
19:30
Glycogen
20:06
Examples: Structures of Polysaccharides
21:42
Let's Draw an (α1→4) & (α1→6) of Amylopectin by Hand.
28:14
More on Glycogen
31:17
Glycogen, Concentration, & The Concept of Osmolarity
35:16
Polysaccharides, Part 2

44m 15s

Intro
0:00
Polysaccharides
0:17
Example: Cellulose
0:34
Glycoside Bond
7:25
Example Illustrations
12:30
Glycosaminoglycans Part 1
15:55
Glycosaminoglycans Part 2
18:34
Glycosaminoglycans & Sulfate Attachments
22:42
β-D-N-Acetylglucosamine
24:49
β-D-N-AcetylGalactosamine
25:42
β-D-Glucuronate
26:44
β-L-Iduronate
27:54
More on Sulfate Attachments
29:49
Hylarunic Acid
32:00
Hyaluronates
39:32
Other Glycosaminoglycans
40:46
Glycoconjugates

44m 23s

Intro
0:00
Glycoconjugates
0:24
Overview
0:25
Proteoglycan
2:53
Glycoprotein
5:20
Glycolipid
7:25
Proteoglycan vs. Glycoprotein
8:15
Cell Surface Diagram
11:17
Proteoglycan Common Structure
14:24
Example: Chondroitin-4-Sulfate
15:06
Glycoproteins
19:50
The Monomers that Commonly Show Up in The Oligo Portions of Glycoproteins
28:02
N-Acetylneuraminic Acid
31:17
L-Furose
32:37
Example of an N-Linked Oligosaccharide
33:21
Cell Membrane Structure
36:35
Glycolipids & Lipopolysaccharide
37:22
Structure Example
41:28
More Example Problems with Carbohydrates

40m 22s

Intro
0:00
Example 1
1:09
Example 2
2:34
Example 3
5:12
Example 4
16:19
Question
16:20
Solution
17:25
Example 5
24:18
Question
24:19
Structure of 2,3-Di-O-Methylglucose
26:47
Part A
28:11
Part B
33:46
Section 7: Lipids
Fatty Acids & Triacylglycerols

54m 55s

Intro
0:00
Fatty Acids
0:32
Lipids Overview
0:34
Introduction to Fatty Acid
3:18
Saturated Fatty Acid
6:13
Unsaturated or Polyunsaturated Fatty Acid
7:07
Saturated Fatty Acid Example
7:46
Unsaturated Fatty Acid Example
9:06
Notation Example: Chain Length, Degree of Unsaturation, & Double Bonds Location of Fatty Acid
11:56
Example 1: Draw the Structure
16:18
Example 2: Give the Shorthand for cis,cis-5,8-Hexadecadienoic Acid
20:12
Example 3
23:12
Solubility of Fatty Acids
25:45
Melting Points of Fatty Acids
29:40
Triacylglycerols
34:13
Definition of Triacylglycerols
34:14
Structure of Triacylglycerols
35:08
Example: Triacylglycerols
40:23
Recall Ester Formation
43:57
The Body's Primary Fuel-Reserves
47:22
Two Primary Advantages to Storing Energy as Triacylglycerols Instead of Glycogen: Number 1
49:24
Two Primary Advantages to Storing Energy as Triacylglycerols Instead of Glycogen: Number 2
51:54
Membrane Lipids

38m 51s

Intro
0:00
Membrane Lipids
0:26
Definition of Membrane Lipids
0:27
Five Major Classes of Membrane Lipids
2:38
Glycerophospholipids
5:04
Glycerophospholipids Overview
5:05
The X Group
8:05
Example: Phosphatidyl Ethanolamine
10:51
Example: Phosphatidyl Choline
13:34
Phosphatidyl Serine
15:16
Head Groups
16:50
Ether Linkages Instead of Ester Linkages
20:05
Galactolipids
23:39
Galactolipids Overview
23:40
Monogalactosyldiacylglycerol: MGDG
25:17
Digalactosyldiacylglycerol: DGDG
28:13
Structure Examples 1: Lipid Bilayer
31:35
Structure Examples 2: Cross Section of a Cell
34:56
Structure Examples 3: MGDG & DGDG
36:28
Membrane Lipids, Part 2

38m 20s

Intro
0:00
Sphingolipids
0:11
Sphingolipid Overview
0:12
Sphingosine Structure
1:42
Ceramide
3:56
Subclasses of Sphingolipids Overview
6:00
Subclasses of Sphingolipids: Sphingomyelins
7:53
Sphingomyelins
7:54
Subclasses of Sphingolipids: Glycosphingolipid
12:47
Glycosphingolipid Overview
12:48
Cerebrosides & Globosides Overview
14:33
Example: Cerebrosides
15:43
Example: Globosides
17:14
Subclasses of Sphingolipids: Gangliosides
19:07
Gangliosides
19:08
Medical Application: Tay-Sachs Disease
23:34
Sterols
30:45
Sterols: Basic Structure
30:46
Important Example: Cholesterol
32:01
Structures Example
34:13
The Biologically Active Lipids

48m 36s

Intro
0:00
The Biologically Active Lipids
0:44
Phosphatidyl Inositol Structure
0:45
Phosphatidyl Inositol Reaction
3:24
Image Example
12:49
Eicosanoids
14:12
Arachidonic Acid & Membrane Lipid Containing Arachidonic Acid
18:41
Three Classes of Eicosanoids
20:42
Overall Structures
21:38
Prostagladins
22:56
Thromboxane
27:19
Leukotrienes
30:19
More On The Biologically Active Lipids
33:34
Steroid Hormones
33:35
Fat Soluble Vitamins
38:25
Vitamin D₃
40:40
Vitamin A
43:17
Vitamin E
45:12
Vitamin K
47:17
Section 8: Energy & Biological Systems (Bioenergetics)
Thermodynamics, Free Energy & Equilibrium

45m 51s

Intro
0:00
Thermodynamics, Free Energy and Equilibrium
1:03
Reaction: Glucose + Pi → Glucose 6-Phosphate
1:50
Thermodynamics & Spontaneous Processes
3:31
In Going From Reactants → Product, a Reaction Wants to Release Heat
6:30
A Reaction Wants to Become More Disordered
9:10
∆H < 0
10:30
∆H > 0
10:57
∆S > 0
11:23
∆S <0
11:56
∆G = ∆H - T∆S at Constant Pressure
12:15
Gibbs Free Energy
15:00
∆G < 0
16:49
∆G > 0
17:07
Reference Frame For Thermodynamics Measurements
17:57
More On BioChemistry Standard
22:36
Spontaneity
25:36
Keq
31:45
Example: Glucose + Pi → Glucose 6-Phosphate
34:14
Example Problem 1
40:25
Question
40:26
Solution
41:12
More on Thermodynamics & Free Energy

37m 6s

Intro
0:00
More on Thermodynamics & Free Energy
0:16
Calculating ∆G Under Standard Conditions
0:17
Calculating ∆G Under Physiological Conditions
2:05
∆G < 0
5:39
∆G = 0
7:03
Reaction Moving Forward Spontaneously
8:00
∆G & The Maximum Theoretical Amount of Free Energy Available
10:36
Example Problem 1
13:11
Reactions That Have Species in Common
17:48
Example Problem 2: Part 1
20:10
Example Problem 2: Part 2- Enzyme Hexokinase & Coupling
25:08
Example Problem 2: Part 3
30:34
Recap
34:45
ATP & Other High-Energy Compounds

44m 32s

Intro
0:00
ATP & Other High-Energy Compounds
0:10
Endergonic Reaction Coupled With Exergonic Reaction
0:11
Major Theme In Metabolism
6:56
Why the ∆G°' for ATP Hydrolysis is Large & Negative
12:24
∆G°' for ATP Hydrolysis
12:25
Reason 1: Electrostatic Repulsion
14:24
Reason 2: Pi & Resonance Forms
15:33
Reason 3: Concentrations of ADP & Pi
17:32
ATP & Other High-Energy Compounds Cont'd
18:48
More On ∆G°' & Hydrolysis
18:49
Other Compounds That Have Large Negative ∆G°' of Hydrolysis: Phosphoenol Pyruvate (PEP)
25:14
Enzyme Pyruvate Kinase
30:36
Another High Energy Molecule: 1,3 Biphosphoglycerate
36:17
Another High Energy Molecule: Phophocreatine
39:41
Phosphoryl Group Transfers

30m 8s

Intro
0:00
Phosphoryl Group Transfer
0:27
Phosphoryl Group Transfer Overview
0:28
Example: Glutamate → Glutamine Part 1
7:11
Example: Glutamate → Glutamine Part 2
13:29
ATP Not Only Transfers Phosphoryl, But Also Pyrophosphoryl & Adenylyl Groups
17:03
Attack At The γ Phosphorous Transfers a Phosphoryl
19:02
Attack At The β Phosphorous Gives Pyrophosphoryl
22:44
Oxidation-Reduction Reactions

49m 46s

Intro
0:00
Oxidation-Reduction Reactions
1:32
Redox Reactions
1:33
Example 1: Mg + Al³⁺ → Mg²⁺ + Al
3:49
Reduction Potential Definition
10:47
Reduction Potential Example
13:38
Organic Example
22:23
Review: How To Find The Oxidation States For Carbon
24:15
Examples: Oxidation States For Carbon
27:45
Example 1: Oxidation States For Carbon
27:46
Example 2: Oxidation States For Carbon
28:36
Example 3: Oxidation States For Carbon
29:18
Example 4: Oxidation States For Carbon
29:44
Example 5: Oxidation States For Carbon
30:10
Example 6: Oxidation States For Carbon
30:40
Example 7: Oxidation States For Carbon
31:20
Example 8: Oxidation States For Carbon
32:10
Example 9: Oxidation States For Carbon
32:52
Oxidation-Reduction Reactions, cont'd
35:22
More On Reduction Potential
35:28
Lets' Start With ∆G = ∆G°' + RTlnQ
38:29
Example: Oxidation Reduction Reactions
41:42
More On Oxidation-Reduction Reactions

56m 34s

Intro
0:00
More On Oxidation-Reduction Reactions
0:10
Example 1: What If the Concentrations Are Not Standard?
0:11
Alternate Procedure That Uses The 1/2 Reactions Individually
8:57
Universal Electron Carriers in Aqueous Medium: NAD+ & NADH
15:12
The Others Are…
19:22
NAD+ & NADP Coenzymes
20:56
FMN & FAD
22:03
Nicotinamide Adenine Dinucleotide (Phosphate)
23:03
Reduction 1/2 Reactions
36:10
Ratio of NAD+ : NADH
36:52
Ratio of NADPH : NADP+
38:02
Specialized Roles of NAD+ & NADPH
38:48
Oxidoreductase Enzyme Overview
40:26
Examples of Oxidoreductase
43:32
The Flavin Nucleotides
46:46
Example Problems For Bioenergetics

42m 12s

Intro
0:00
Example 1: Calculate the ∆G°' For The Following Reaction
1:04
Example 1: Question
1:05
Example 1: Solution
2:20
Example 2: Calculate the Keq For the Following
4:20
Example 2: Question
4:21
Example 2: Solution
5:54
Example 3: Calculate the ∆G°' For The Hydrolysis of ATP At 25°C
8:52
Example 3: Question
8:53
Example 3: Solution
10:30
Example 3: Alternate Procedure
13:48
Example 4: Problems For Bioenergetics
16:46
Example 4: Questions
16:47
Example 4: Part A Solution
21:19
Example 4: Part B Solution
23:26
Example 4: Part C Solution
26:12
Example 5: Problems For Bioenergetics
29:27
Example 5: Questions
29:35
Example 5: Solution - Part 1
32:16
Example 5: Solution - Part 2
34:39
Section 9: Glycolysis and Gluconeogenesis
Overview of Glycolysis I

43m 32s

Intro
0:00
Overview of Glycolysis
0:48
Three Primary Paths For Glucose
1:04
Preparatory Phase of Glycolysis
4:40
Payoff Phase of Glycolysis
6:40
Glycolysis Reactions Diagram
7:58
Enzymes of Glycolysis
12:41
Glycolysis Reactions
16:02
Step 1
16:03
Step 2
18:03
Step 3
18:52
Step 4
20:08
Step 5
21:42
Step 6
22:44
Step 7
24:22
Step 8
25:11
Step 9
26:00
Step 10
26:51
Overview of Glycolysis Cont.
27:28
The Overall Reaction for Glycolysis
27:29
Recall The High-Energy Phosphorylated Compounds Discusses In The Bioenergetics Unit
33:10
What Happens To The Pyruvate That Is Formed?
37:58
Glycolysis II

1h 1m 47s

Intro
0:00
Glycolysis Step 1: The Phosphorylation of Glucose
0:27
Glycolysis Step 1: Reaction
0:28
Hexokinase
2:28
Glycolysis Step 1: Mechanism-Simple Nucleophilic Substitution
6:34
Glycolysis Step 2: Conversion of Glucose 6-Phosphate → Fructose 6-Phosphate
11:33
Glycolysis Step 2: Reaction
11:34
Glycolysis Step 2: Mechanism, Part 1
14:40
Glycolysis Step 2: Mechanism, Part 2
18:16
Glycolysis Step 2: Mechanism, Part 3
19:56
Glycolysis Step 2: Mechanism, Part 4 (Ring Closing & Dissociation)
21:54
Glycolysis Step 3: Conversion of Fructose 6-Phosphate to Fructose 1,6-Biphosphate
24:16
Glycolysis Step 3: Reaction
24:17
Glycolysis Step 3: Mechanism
26:40
Glycolysis Step 4: Cleavage of Fructose 1,6-Biphosphate
31:10
Glycolysis Step 4: Reaction
31:11
Glycolysis Step 4: Mechanism, Part 1 (Binding & Ring Opening)
35:26
Glycolysis Step 4: Mechanism, Part 2
37:40
Glycolysis Step 4: Mechanism, Part 3
39:30
Glycolysis Step 4: Mechanism, Part 4
44:00
Glycolysis Step 4: Mechanism, Part 5
46:34
Glycolysis Step 4: Mechanism, Part 6
49:00
Glycolysis Step 4: Mechanism, Part 7
50:12
Hydrolysis of The Imine
52:33
Glycolysis Step 5: Conversion of Dihydroxyaceton Phosphate to Glyceraldehyde 3-Phosphate
55:38
Glycolysis Step 5: Reaction
55:39
Breakdown and Numbering of Sugar
57:40
Glycolysis III

59m 17s

Intro
0:00
Glycolysis Step 5: Conversion of Dihydroxyaceton Phosphate to Glyceraldehyde 3-Phosphate
0:44
Glycolysis Step 5: Mechanism, Part 1
0:45
Glycolysis Step 5: Mechanism, Part 2
3:53
Glycolysis Step 6: Oxidation of Glyceraldehyde 3-Phosphate to 1,3-Biphosphoglycerate
5:14
Glycolysis Step 6: Reaction
5:15
Glycolysis Step 6: Mechanism, Part 1
8:52
Glycolysis Step 6: Mechanism, Part 2
12:58
Glycolysis Step 6: Mechanism, Part 3
14:26
Glycolysis Step 6: Mechanism, Part 4
16:23
Glycolysis Step 7: Phosphoryl Transfer From 1,3-Biphosphoglycerate to ADP to Form ATP
19:08
Glycolysis Step 7: Reaction
19:09
Substrate-Level Phosphorylation
23:18
Glycolysis Step 7: Mechanism (Nucleophilic Substitution)
26:57
Glycolysis Step 8: Conversion of 3-Phosphoglycerate to 2-Phosphoglycerate
28:44
Glycolysis Step 8: Reaction
28:45
Glycolysis Step 8: Mechanism, Part 1
30:08
Glycolysis Step 8: Mechanism, Part 2
32:24
Glycolysis Step 8: Mechanism, Part 3
34:02
Catalytic Cycle
35:42
Glycolysis Step 9: Dehydration of 2-Phosphoglycerate to Phosphoenol Pyruvate
37:20
Glycolysis Step 9: Reaction
37:21
Glycolysis Step 9: Mechanism, Part 1
40:12
Glycolysis Step 9: Mechanism, Part 2
42:01
Glycolysis Step 9: Mechanism, Part 3
43:58
Glycolysis Step 10: Transfer of a Phosphoryl Group From Phosphoenol Pyruvate To ADP To Form ATP
45:16
Glycolysis Step 10: Reaction
45:17
Substrate-Level Phosphorylation
48:32
Energy Coupling Reaction
51:24
Glycolysis Balance Sheet
54:15
Glycolysis Balance Sheet
54:16
What Happens to The 6 Carbons of Glucose?
56:22
What Happens to 2 ADP & 2 Pi?
57:04
What Happens to The 4e⁻ ?
57:15
Glycolysis IV

39m 47s

Intro
0:00
Feeder Pathways
0:42
Feeder Pathways Overview
0:43
Starch, Glycogen
2:25
Lactose
4:38
Galactose
4:58
Manose
5:22
Trehalose
5:45
Sucrose
5:56
Fructose
6:07
Fates of Pyruvate: Aerobic & Anaerobic Conditions
7:39
Aerobic Conditions & Pyruvate
7:40
Anaerobic Fates of Pyruvate
11:18
Fates of Pyruvate: Lactate Acid Fermentation
14:10
Lactate Acid Fermentation
14:11
Fates of Pyruvate: Ethanol Fermentation
19:01
Ethanol Fermentation Reaction
19:02
TPP: Thiamine Pyrophosphate (Functions and Structure)
23:10
Ethanol Fermentation Mechanism, Part 1
27:53
Ethanol Fermentation Mechanism, Part 2
29:06
Ethanol Fermentation Mechanism, Part 3
31:15
Ethanol Fermentation Mechanism, Part 4
32:44
Ethanol Fermentation Mechanism, Part 5
34:33
Ethanol Fermentation Mechanism, Part 6
35:48
Gluconeogenesis I

41m 34s

Intro
0:00
Gluconeogenesis, Part 1
1:02
Gluconeogenesis Overview
1:03
3 Glycolytic Reactions That Are Irreversible Under Physiological Conditions
2:29
Gluconeogenesis Reactions Overview
6:17
Reaction: Pyruvate to Oxaloacetate
11:07
Reaction: Oxaloacetate to Phosphoenolpyruvate (PEP)
13:29
First Pathway That Pyruvate Can Take to Become Phosphoenolpyruvate
15:24
Second Pathway That Pyruvate Can Take to Become Phosphoenolpyruvate
21:00
Transportation of Pyruvate From The Cytosol to The Mitochondria
24:15
Transportation Mechanism, Part 1
26:41
Transportation Mechanism, Part 2
30:43
Transportation Mechanism, Part 3
34:04
Transportation Mechanism, Part 4
38:14
Gluconeogenesis II

34m 18s

Intro
0:00
Oxaloacetate → Phosphoenolpyruvate (PEP)
0:35
Mitochondrial Membrane Does Not Have a Transporter for Oxaloactate
0:36
Reaction: Oxaloacetate to Phosphoenolpyruvate (PEP)
3:36
Mechanism: Oxaloacetate to Phosphoenolpyruvate (PEP)
4:48
Overall Reaction: Pyruvate to Phosphoenolpyruvate
7:01
Recall The Two Pathways That Pyruvate Can Take to Become Phosphoenolpyruvate
10:16
NADH in Gluconeogenesis
12:29
Second Pathway: Lactate → Pyruvate
18:22
Cytosolic PEP Carboxykinase, Mitochondrial PEP Carboxykinase, & Isozymes
18:23
2nd Bypass Reaction
23:04
3rd Bypass Reaction
24:01
Overall Process
25:17
Other Feeder Pathways For Gluconeogenesis
26:35
Carbon Intermediates of The Citric Acid Cycle
26:36
Amino Acids & The Gluconeogenic Pathway
29:45
Glycolysis & Gluconeogenesis Are Reciprocally Regulated
32:00
The Pentose Phosphate Pathway

42m 52s

Intro
0:00
The Pentose Phosphate Pathway Overview
0:17
The Major Fate of Glucose-6-Phosphate
0:18
The Pentose Phosphate Pathway (PPP) Overview
1:00
Oxidative Phase of The Pentose Phosphate Pathway
4:33
Oxidative Phase of The Pentose Phosphate Pathway: Reaction Overview
4:34
Ribose-5-Phosphate: Glutathione & Reductive Biosynthesis
9:02
Glucose-6-Phosphate to 6-Phosphogluconate
12:48
6-Phosphogluconate to Ribulose-5-Phosphate
15:39
Ribulose-5-Phosphate to Ribose-5-Phosphate
17:05
Non-Oxidative Phase of The Pentose Phosphate Pathway
19:55
Non-Oxidative Phase of The Pentose Phosphate Pathway: Overview
19:56
General Transketolase Reaction
29:03
Transaldolase Reaction
35:10
Final Transketolase Reaction
39:10
Section 10: The Citric Acid Cycle (Krebs Cycle)
Citric Acid Cycle I

36m 10s

Intro
0:00
Stages of Cellular Respiration
0:23
Stages of Cellular Respiration
0:24
From Pyruvate to Acetyl-CoA
6:56
From Pyruvate to Acetyl-CoA: Pyruvate Dehydrogenase Complex
6:57
Overall Reaction
8:42
Oxidative Decarboxylation
11:54
Pyruvate Dehydrogenase (PDH) & Enzymes
15:30
Pyruvate Dehydrogenase (PDH) Requires 5 Coenzymes
17:15
Molecule of CoEnzyme A
18:52
Thioesters
20:56
Lipoic Acid
22:31
Lipoate Is Attached To a Lysine Residue On E₂
24:42
Pyruvate Dehydrogenase Complex: Reactions
26:36
E1: Reaction 1 & 2
30:38
E2: Reaction 3
31:58
E3: Reaction 4 & 5
32:44
Substrate Channeling
34:17
Citric Acid Cycle II

49m 20s

Intro
0:00
Citric Acid Cycle Reactions Overview
0:26
Citric Acid Cycle Reactions Overview: Part 1
0:27
Citric Acid Cycle Reactions Overview: Part 2
7:03
Things to Note
10:58
Citric Acid Cycle Reactions & Mechanism
13:57
Reaction 1: Formation of Citrate
13:58
Reaction 1: Mechanism
19:01
Reaction 2: Citrate to Cis Aconistate to Isocitrate
28:50
Reaction 3: Isocitrate to α-Ketoglutarate
32:35
Reaction 3: Two Isocitrate Dehydrogenase Enzymes
36:24
Reaction 3: Mechanism
37:33
Reaction 4: Oxidation of α-Ketoglutarate to Succinyl-CoA
41:38
Reaction 4: Notes
46:34
Citric Acid Cycle III

44m 11s

Intro
0:00
Citric Acid Cycle Reactions & Mechanism
0:21
Reaction 5: Succinyl-CoA to Succinate
0:24
Reaction 5: Reaction Sequence
2:35
Reaction 6: Oxidation of Succinate to Fumarate
8:28
Reaction 7: Fumarate to Malate
10:17
Reaction 8: Oxidation of L-Malate to Oxaloacetate
14:15
More On The Citric Acid Cycle
17:17
Energy from Oxidation
17:18
How Can We Transfer This NADH Into the Mitochondria
27:10
Citric Cycle is Amphibolic - Works In Both Anabolic & Catabolic Pathways
32:06
Biosynthetic Processes
34:29
Anaplerotic Reactions Overview
37:26
Anaplerotic: Reaction 1
41:42
Section 11: Catabolism of Fatty Acids
Fatty Acid Catabolism I

48m 11s

Intro
0:00
Introduction to Fatty Acid Catabolism
0:21
Introduction to Fatty Acid Catabolism
0:22
Vertebrate Cells Obtain Fatty Acids for Catabolism From 3 Sources
2:16
Diet: Part 1
4:00
Diet: Part 2
5:35
Diet: Part 3
6:20
Diet: Part 4
6:47
Diet: Part 5
10:18
Diet: Part 6
10:54
Diet: Part 7
12:04
Diet: Part 8
12:26
Fats Stored in Adipocytes Overview
13:54
Fats Stored in Adipocytes (Fat Cells): Part 1
16:13
Fats Stored in Adipocytes (Fat Cells): Part 2
17:16
Fats Stored in Adipocytes (Fat Cells): Part 3
19:42
Fats Stored in Adipocytes (Fat Cells): Part 4
20:52
Fats Stored in Adipocytes (Fat Cells): Part 5
22:56
Mobilization of TAGs Stored in Fat Cells
24:35
Fatty Acid Oxidation
28:29
Fatty Acid Oxidation
28:48
3 Reactions of the Carnitine Shuttle
30:42
Carnitine Shuttle & The Mitochondrial Matrix
36:25
CAT I
43:58
Carnitine Shuttle is the Rate-Limiting Steps
46:24
Fatty Acid Catabolism II

45m 58s

Intro
0:00
Fatty Acid Catabolism
0:15
Fatty Acid Oxidation Takes Place in 3 Stages
0:16
β-Oxidation
2:05
β-Oxidation Overview
2:06
Reaction 1
4:20
Reaction 2
7:35
Reaction 3
8:52
Reaction 4
10:16
β-Oxidation Reactions Discussion
11:34
Notes On β-Oxidation
15:14
Double Bond After The First Reaction
15:15
Reaction 1 is Catalyzed by 3 Isozymes of Acyl-CoA Dehydrogenase
16:04
Reaction 2 & The Addition of H₂O
18:38
After Reaction 4
19:24
Production of ATP
20:04
β-Oxidation of Unsaturated Fatty Acid
21:25
β-Oxidation of Unsaturated Fatty Acid
22:36
β-Oxidation of Mono-Unsaturates
24:49
β-Oxidation of Mono-Unsaturates: Reaction 1
24:50
β-Oxidation of Mono-Unsaturates: Reaction 2
28:43
β-Oxidation of Mono-Unsaturates: Reaction 3
30:50
β-Oxidation of Mono-Unsaturates: Reaction 4
31:06
β-Oxidation of Polyunsaturates
32:29
β-Oxidation of Polyunsaturates: Part 1
32:30
β-Oxidation of Polyunsaturates: Part 2
37:08
β-Oxidation of Polyunsaturates: Part 3
40:25
Fatty Acid Catabolism III

33m 18s

Intro
0:00
Fatty Acid Catabolism
0:43
Oxidation of Fatty Acids With an Odd Number of Carbons
0:44
β-oxidation in the Mitochondrion & Two Other Pathways
9:08
ω-oxidation
10:37
α-oxidation
17:22
Ketone Bodies
19:08
Two Fates of Acetyl-CoA Formed by β-Oxidation Overview
19:09
Ketone Bodies: Acetone
20:42
Ketone Bodies: Acetoacetate
20:57
Ketone Bodies: D-β-hydroxybutyrate
21:25
Two Fates of Acetyl-CoA Formed by β-Oxidation: Part 1
22:05
Two Fates of Acetyl-CoA Formed by β-Oxidation: Part 2
26:59
Two Fates of Acetyl-CoA Formed by β-Oxidation: Part 3
30:52
Section 12: Catabolism of Amino Acids and the Urea Cycle
Overview & The Aminotransferase Reaction

40m 59s

Intro
0:00
Overview of The Aminotransferase Reaction
0:25
Overview of The Aminotransferase Reaction
0:26
The Aminotransferase Reaction: Process 1
3:06
The Aminotransferase Reaction: Process 2
6:46
Alanine From Muscle Tissue
10:54
Bigger Picture of the Aminotransferase Reaction
14:52
Looking Closely at Process 1
19:04
Pyridoxal Phosphate (PLP)
24:32
Pyridoxamine Phosphate
25:29
Pyridoxine (B6)
26:38
The Function of PLP
27:12
Mechanism Examples
28:46
Reverse Reaction: Glutamate to α-Ketoglutarate
35:34
Glutamine & Alanine: The Urea Cycle I

39m 18s

Intro
0:00
Glutamine & Alanine: The Urea Cycle I
0:45
Excess Ammonia, Glutamate, and Glutamine
0:46
Glucose-Alanine Cycle
9:54
Introduction to the Urea Cycle
20:56
The Urea Cycle: Production of the Carbamoyl Phosphate
22:59
The Urea Cycle: Reaction & Mechanism Involving the Carbamoyl Phosphate Synthetase
33:36
Glutamine & Alanine: The Urea Cycle II

36m 21s

Intro
0:00
Glutamine & Alanine: The Urea Cycle II
0:14
The Urea Cycle Overview
0:34
Reaction 1: Ornithine → Citrulline
7:30
Reaction 2: Citrulline → Citrullyl-AMP
11:15
Reaction 2': Citrullyl-AMP → Argininosuccinate
15:25
Reaction 3: Argininosuccinate → Arginine
20:42
Reaction 4: Arginine → Orthinine
24:00
Links Between the Citric Acid Cycle & the Urea Cycle
27:47
Aspartate-argininosuccinate Shunt
32:36
Amino Acid Catabolism

47m 58s

Intro
0:00
Amino Acid Catabolism
0:10
Common Amino Acids and 6 Major Products
0:11
Ketogenic Amino Acid
1:52
Glucogenic Amino Acid
2:51
Amino Acid Catabolism Diagram
4:18
Cofactors That Play a Role in Amino Acid Catabolism
7:00
Biotin
8:42
Tetrahydrofolate
10:44
S-Adenosylmethionine (AdoMet)
12:46
Tetrahydrobiopterin
13:53
S-Adenosylmethionine & Tetrahydrobiopterin Molecules
14:41
Catabolism of Phenylalanine
18:30
Reaction 1: Phenylalanine to Tyrosine
18:31
Reaction 2: Tyrosine to p-Hydroxyphenylpyruvate
21:36
Reaction 3: p-Hydroxyphenylpyruvate to Homogentisate
23:50
Reaction 4: Homogentisate to Maleylacetoacetate
25:42
Reaction 5: Maleylacetoacetate to Fumarylacetoacetate
28:20
Reaction 6: Fumarylacetoacetate to Fumarate & Succinyl-CoA
29:51
Reaction 7: Fate of Fumarate & Succinyl-CoA
31:14
Phenylalanine Hydroxylase
33:33
The Phenylalanine Hydroxylase Reaction
33:34
Mixed-Function Oxidases
40:26
When Phenylalanine Hydoxylase is Defective: Phenylketonuria (PKU)
44:13
Section 13: Oxidative Phosphorylation and ATP Synthesis
Oxidative Phosphorylation I

41m 11s

Intro
0:00
Oxidative Phosphorylation
0:54
Oxidative Phosphorylation Overview
0:55
Mitochondrial Electron Transport Chain Diagram
7:15
Enzyme Complex I of the Electron Transport Chain
12:27
Enzyme Complex II of the Electron Transport Chain
14:02
Enzyme Complex III of the Electron Transport Chain
14:34
Enzyme Complex IV of the Electron Transport Chain
15:30
Complexes Diagram
16:25
Complex I
18:25
Complex I Overview
18:26
What is Ubiquinone or Coenzyme Q?
20:02
Coenzyme Q Transformation
22:37
Complex I Diagram
24:47
Fe-S Proteins
26:42
Transfer of H⁺
29:42
Complex II
31:06
Succinate Dehydrogenase
31:07
Complex II Diagram & Process
32:54
Other Substrates Pass Their e⁻ to Q: Glycerol 3-Phosphate
37:31
Other Substrates Pass Their e⁻ to Q: Fatty Acyl-CoA
39:02
Oxidative Phosphorylation II

36m 27s

Intro
0:00
Complex III
0:19
Complex III Overview
0:20
Complex III: Step 1
1:56
Complex III: Step 2
6:14
Complex IV
8:42
Complex IV: Cytochrome Oxidase
8:43
Oxidative Phosphorylation, cont'd
17:18
Oxidative Phosphorylation: Summary
17:19
Equation 1
19:13
How Exergonic is the Reaction?
21:03
Potential Energy Represented by Transported H⁺
27:24
Free Energy Change for the Production of an Electrochemical Gradient Via an Ion Pump
28:48
Free Energy Change in Active Mitochondria
32:02
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Lecture Comments (6)

0 answers

Post by Anthony Villarama on January 4, 2019

Thank you Raffi. You are a world class teacher. I want to be like someday.

1 answer

Last reply by: Professor Hovasapian
Wed Mar 5, 2014 3:46 PM

Post by Billy Jabbar on March 5, 2014

Interesting lecture Dr. Hovasapian.  

My instructor skipped over Edman degradation in class and instead decided to focus on newer Mass Spectrometry techniques that are beginning to replace classical protein sequencing techniques like Edman Degradation. I still thought it was worth learning because I see many other classes do cover it, but was wondering if you may include a lecture on applications of Mass Spectrometry for protein sequencing in the future.  Thanks!

0 answers

Post by Matthew Humes on September 29, 2013

I would like to agree with Omri, very clear and concise. A welcome change from my lectures =)

1 answer

Last reply by: Professor Hovasapian
Fri Sep 20, 2013 11:56 PM

Post by omri shick on September 20, 2013

thank you! it is very helpful lecture !!

Amino Acid Sequencing of a Peptide Chain

Lecture Slides are screen-captured images of important points in the lecture. Students can download and print out these lecture slide images to do practice problems as well as take notes while watching the lecture.

  • Intro 0:00
  • Amino Acid Sequencing of a Peptide Chain 0:30
    • Amino Acid Sequence and Its Structure
    • Edman Degradation: Overview
    • Edman Degradation: Reaction - Part 1
    • Edman Degradation: Reaction - Part 2
    • Edman Degradation: Reaction - Part 3
    • Mechanism Step 1: PTC (Phenylthiocarbamyl) Formation
    • Mechanism Step 2: Ring Formation & Peptide Bond Cleavage
  • Example: Write Out the Edman Degradation for the Tripeptide Ala-Tyr-Ser 30:29
    • Step 1
    • Step 2
    • Step 3
    • Step 4
    • Step 5
    • Step 6

Transcription: Amino Acid Sequencing of a Peptide Chain

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

At the close of the last lesson, we talked about the levels of protein structure; we had primary, secondary, tertiary and quaternary.0004

Today, we're going to talk about the primary structure- the amino acid sequence.0012

We want to know what is the sequence of amino acids- which is next to which, which amino acid is next to which, and how are they arranged in a linear fashion.0017

That is what we're going to be working on.0028

Let's get started.0029

OK.0032

The amino acid sequence determines how the peptide is going to actually fold, how the peptide will fold, and thus, ultimately determines its structure.0034

As you can see, the primary sequence of a protein, of a peptide, is very, very important because depending on what amino acids are aware, it's going to basically guide how the protein is going to assume its 3-dimensional shape; and it is that 3-dimensional shape which is going to determine its structure and function.0080

Let me write these words a little bit better.0109

The amino acid sequence determines how the peptide will fold and thus ultimately determines its structure and function.0113

Amino acid sequence implies the function, and that's what is important in a protein, what does it do.0126

OK.0135

Well, there are many techniques for elucidating amino acid sequence.0137

We will discuss a chemical method.0160

We will discuss a chemical method still used in laboratories.0165

It is called the Edman degradation.0174

Excuse me.0186

Basically, what the Edman degradation does is it labels and removes the N-terminal amino acid for identification.0187

It labels it for identification.0210

It removes it so that it can be separated, and that way you can identify it.0211

The remaining peptide, now, has a new N-terminal amino acid; and now, what we do is we just repeat the process.0218

That's it.0243

Excuse me.0245

We're basically taking an amino acid and we're labeling the end, cutting it off, identifying it.0247

Next one, labeling the end, cutting it off, identifying it, and we just go down the list.0250

We are just chopping it up until we finally get to the last amino acid.0254

That's all the Edman degradation does, and, of course, this is an automated procedure because we have really, really good chemical control; so we can just put our sample into a machine, and it will do everything for us, and it will give us a read out at the end.0259

It is really quite wonderful.0271

OK.0275

I'm going to do a schematic representation of the Edman degradation describing each step, and then we’ll go ahead and do an example of an Edman degradation with a specific amino acid.0276

OK.0289

I wonder if I should start on a new - yes - let me go ahead and start on a new page here.0291

Yes, that's fine.0296

This is going to be the Edman degradation; let me go ahead and do this in blue.0299

OK.0306

I have to warn you there is going to be a lot of chemical names being thrown around, and there is going to be a lot of chemical structures being thrown around.0307

This is where you have to be really, really, really careful, and that includes me.0315

So, please, by all means, you definitely want to confirm that I'm actually drawing the right structures.0320

I would definitely encourage you to take a look at the Edman degradation procedure in your book to see what they have to say about the particular mechanism and how they draw it- really, really important.0330

But again, ultimately, it is just not about passive learning.0340

You don't just want to look at a diagram and say I understand it; you need to be able to reproduce it.0344

That's when you actually understand it.0350

OK, so, the Edman degradation.0352

Let's start off with just a generic peptide.0354

We have H3, N, C, C, and I'm just going to go ahead and write peptide for the other because again, we're just going to be concerned with the N-terminal, the one on the left.0360

We have the carbonyl carbon there, and we have our R-group attached to the alpha-carbon, and this is A+.0372

The first step is...where should I write this, I'll go ahead and write it here, wonder if I should do it in, this one I'm going to do in black, I think.0379

OK.0402

I'm going to be drawing this thing, N, double bond C, double bond S.0403

OK.0410

What we do is we take this peptide and we react it with something called phenyl isothiocyanate under mildly alkaline conditions.0411

OK.0418

That's the first step0419

Let me go ahead and write this as one.0421

I'm going to write the products below instead of to the right.0425

I'm going to write the steps over here; I just wanted to do it in a schematic way.0427

You know what I need a little bit more room to write this out.0438

One, phenyl isothiocyanate- that is this molecule right here.0443

OK.0454

It is abbreviated PITC, phenyl isothiocyanate, under mild basic conditions, alkaline conditions - there we go - under mild OH.0455

This is the Edman reagent, so you'll often hear it.0471

They might say PITC, or they will just say “use Edman reagent”.0475

This is our Edman reagent; let me go ahead and put that there.0480

This is called the Edman reagent; let me go back to black.0485

OK.0490

When this reaction actually takes place, what you end up with is this product.0491

Let me see.0499

It is going to be this here; let's go ahead and put the H on there.0500

It is going to be C, double bonded S, and it is going to be attached to the N, C, C.0510

This is carbonyl, and this is our peptide.0518

This is our R-group, and we have our H.0523

OK.0527

The bond is formed between this carbon and that nitrogen.0531

This is the bond that is formed- right there.0539

OK.0542

Now, again, let's keep track of our peptide.0544

Our peptide is right here- N, C, C.0548

That is what you want to look for.0550

When you're doing these yourself, again, keep track of your peptide; and you can keep track of it by looking for that N, C, C motif.0551

N to the left, C to the right, C to the right, carbonyl on the second C, R-group on the first C, counting from left to right- here is our peptide, I'm sorry, here is our amino acid.0559

This is the one that we're actually pulling off.0574

This is the isothiocyanate part here.0576

So, what I've actually ended up forming here is something called phenylthiocarbamoyl that refers to this particular arrangement of atoms.0580

Phenyl is this, thio is the sulfur, carbamoyl is this carbon attached to a nitrogen and a nitrogen here.0599

What I've done is I've taken this peptide that I have, and I've created a phenylthiocarbamoyl derivative of it, by reacting it with the phenyl isothiocyanate, the PITC.0606

This phenylthiocarbamoyl derivative, they call it PTC.0620

OK.0626

Now, we'll go to our second step.0627

Now, we'll go ahead and do another black here.0628

OK.0633

This one, we are going to react it with C, COOH; and this is going to be anhydrous.0634

What we're going to do is, we're going to react this PTC with anhydrous trifluoroacetic acid.0644

It is just a weak acid that happens to be a little bit stronger than acidic acid.0663

Actually, any acid will do; it's fine.0667

It just needs to be anhydrous.0669

Now, what happens when this reaction takes place is the following.0672

What you end up with is the following 2 molecules; this is the one that actually breaks the bond that we are trying to break.0675

OK.0683

And, I'll tell you which bond in just a minute once I draw it out.0685

Let' me see.0688

That's fine; I guess I can fit it in here.0689

Let me go back to blue.0691

We have our C, we have our NH, and we have our phenyl group, C, then we have our...here's our N, here is our C, and here's our C, that is our that, and then we have our S, and C, and we have our 1.0693

Let me make sure I have everything on here, N, trivalent, S.0728

OK.0733

Everything is good.0734

Yes, and, of course, we have that plus our new peptide, 3 peptide- the new amino-terminus.0735

The bond that we have actually broken is the following.0750

We've broken this bond; let me do this in black.0754

We have broken this bond, and again I'm going to go through the mechanism in just a little bit, but I just wanted you to see chemically what happens.0765

This thing, when we form this species or again, let me see, N, C, C, keep track of the N, C, C.0771

This is our amino acid.0781

OK.0784

This is our that; this is our that.0785

That is what this is.0787

We want to keep track of our amino acids.0788

This is called, in case you want to know, it's called an anilinothiazolinone.0790

Anilino refers to the phenyl group attached to nitrogen; thiozolinone happens to be this thing, the C, the N, the S, arranged in a ring.0803

OK.0813

Now, we take the third step - oops this is not...this is 2, not number 1, number 1 was that, sorry about that - this is step 2 of the Edman degradation.0816

Now, we're going to go to step 3 of the Edman degradation.0829

Let me move on to the next page, and let me write the molecule in blue.0832

Now, we've pulled off that other peptide.0837

We have that one N-terminus that we've actually broken off; that's the thing that we're going to react.0842

Let me redraw that one; let me draw it here.0847

C, we have NH, we have that, we have N, we have C, we have C, we have S, this is our R group, this is our carbonyl.0852

Let me see; am I missing anything here?0871

No, I don't think so; everything looks good.0874

OK.0877

Now, again, let me, N, C, C, just to keep track of our amino acid or N-terminal.0878

Now, the third step here, what we do is we're just going to react this particular molecule with aqueous acid.0888

So, step 3 is aqueous acid, and what you end up with is the following molecule.0901

Let me do this in...yes, that's fine; I'll go ahead and do it in blue.0913

We have C, we have S, we have N, we have C, we have C, and we have N, and we have phenyl, and we have an H, we have our R group, and we have that.0917

OK.0943

The reason we actually do this step is this thing is more stable than this thing, so it allows us to deal with it better.0946

This is more stable, and it is called phenylthiohydantoin; and this is PTH- that's the acronym.0952

Now, it's ready for identification.0972

There you go.0983

And now, let's go ahead and red N, C, C.0985

That is our motif; that’s what we want to keep track of.0990

All that has happened here is that this thing under acidic conditions, aqueous acidic conditions, has actually rearranged, and has formed something more stable.0993

You want to take a look what has happened form here to here.1004

The only thing that has happened is this carbon right here that is attached to nitrogen, this S, went up to where the nitrogen was, double bond; this nitrogen with the phenyl group came down to where the S was.1006

This S and this thing switched places; that's all that happened- the rearrangement.1019

OK.1026

And, of course, the last part, since now you have the H3N - oops - you have the peptide left over, the fourth step.1028

Let me do this in blue.1038

H3, N+, now, you have the rest of the peptide with a new N-terminal amino acid group.1040

So, step 4, just repeat the process.1047

That is the Edman degradation.1052

The first step is phenyl isothiocyanate, and then after that, you're going to treat it with trifluoroacetic acid; third step, you're going to treat it with aqueous acid, and you're going to form this molecule right here- this phenylthiohydantoin.1054

You've basically taken this N-amino acid, and you've labeled it with this thing.1075

You've made a derivative of this thing for that thing, and now, you can identify it; and you just repeat the process, go down the chain.1080

OK.1087

Now, let's take a look at some mechanisms.1088

It's important to talk about mechanisms, how electrons move, arrow pushing.1090

You remember from organic chemistry, electrons go this way, nucleophile, electrophile.1095

If it is something that's strange to you or perhaps you’re not too familiar with it, it intimidates you a little bit, don't worry about it.1101

I think it will just be reasonably clear what are these that's going on.1108

Don't attach any more deeper meaning than what it actually is.1111

It is just electrons moving around forming bonds.1116

You remember in general chemistry, we just sort of do this chemistry, and we wouldn’t talk about how it happened.1119

When you got to organic chemistry, that's when you started talking about "OK, this carbon is moving in here, these electrons are forming this bond, this bond is breaking"- that's all a mechanism is.1124

It is a molecular level, single step, what's happening.1134

OK.1140

Let's see if we can do - let's do this in blue - mechanism for PTC formation.1141

That is the phenylthiocarbamoyl formation.1156

This is step one.1167

OK.1175

Here we go.1176

Let's go ahead and draw out the phenyl isothiocyanate first.1177

So, I’m going to draw this vertically.1182

Actually, let me do this in black.1185

OK.1188

And, we've got N, we have C and S, so this is our PITC- phenyl isothiocyanate.1191

Now, let's go ahead and write our H2; this is N.1200

I’ll go ahead and put the electrons on the nitrogen; I’ve got N, C, C, and then I've got...I'm going to actually write out the second, N, C, C, N, C, C.1204

Carbonyl goes here; carbonyl goes here.1220

I'm just going to do it for a dipeptide.1222

This is going to be the R1 group; this is going to be the R2 group, and let's go ahead and put an H on that nitrogen.1225

OK.1232

Here is what happens.1233

These electrons, it is a nucleophile; this nitrogen is a nucleophile.1236

This carbon here that is attached to nitrogen and sulfur, it's the electrophile.1242

It is a little bit positively charged.1248

Nitrogen is an electronegative element.1249

It's going to pull electrons away from that.1252

This is negatively charged.1253

So, these electrons are going to attack here, and when these electrons come in, electrons that are there have to make room for these that come in, so they have to go away.1256

These electrons move away and they grab an H from the solution; and what you end up getting is the following.1269

Should I draw it?1280

Yes, that is fine; I'll go ahead and draw it horizontally.1284

N, H, C, double bond S, this is N, H.1292

It is going to be C, C, and then N, C, C.1302

OK.1311

Well, that's fine; I'll do this in just a minute.1312

N, C, C, this is the carbonyl, this is our R1 group, there is an H here, this is our R2 group.1315

So, the bond that we formed is this bond right here.1325

That is the bond that we formed.1329

These electrons formed this bond.1330

Now, notice, it has 2 hydrogens on it, but this nitrogen now, has 1 hydrogen on it.1333

So, I'm going to go ahead and write this minus H plus.1339

That means that it has given up that hydrogen.1342

Once this bond forms, now, nitrogen has 1, 2, 3, 4 things attached to it.1343

It is going to be positively charged.1351

It is going to release that hydrogen in the solution.1352

This is the mechanism.1357

Nitrogen is the nucleophile; this carbon of the PITC is the electrophile- standard, basic mechanism, single step.1359

OK.1369

This is our PTC, phenylthiocarbamoyl.1370

OK.1377

Now, let's go ahead and do the mechanism for the second step.1380

This is very important.1384

This one we'll do in blue again.1386

This is going to be the mechanism for ring formation and peptide bond cleavage.1389

This is the big one.1408

This is step 2.1414

This is where we add the trifluoroacetic acid.1416

OK, step 2.1419

So, we've added the trifluoroacetic acid, this is what happens.1421

Let's draw our molecule again, and we'll make sure to draw it very, very carefully.1424

And again, you need to be able to reproduce this.1429

OK.1433

It's the only way you'll have a full grasp of what it is that is going on.1434

We have N, C, C, N, C, C.1437

There is an H here, our carbonyl goes there, carbonyl goes there.1445

This is our R1 group; this is our R2 group, and let me go ahead and put the electrons on the nitrogen on that one.1450

OK.1462

Now, here is what happens.1463

Alright.1466

We are just going to be pushing arrows; electrons are going to be moving around.1467

Here is what happens.1471

Actually, you know what, I'm going to make this arrow a little bit smaller here because I want to do 3 structures on this page.1473

OK.1481

These arrows right here on the nitrogen, that is N, C, C, our N-terminal, OK, this nitrogen.1482

Again, look for the N, C, C.1489

This is N, C, N; that's not it- N, C, C.1491

This is your terminal amino acid.1493

These electrons, they go that way.1498

These electrons, they push, these electrons they attack that.1501

You know what, I'm going to do this in a different color.1509

Sorry about that.1513

Let me do this in red.1516

These electrons go down here to form a double bond.1520

They push these electrons; they attack the carbonyl right here, and these actually end up going up onto oxygen.1522

So, what you end up getting is this tetrahedral intermediate, which is very typical of carbonyl reactivity.1533

What you end up with is the following.1540

Now, I'm going to retain certain structural features.1542

I'm going to keep this C, this arrangement, while I draw a structure.1545

It's going to be...let me do this in blue.1551

I'll try to do it underneath.1554

C, C, O-, N, C, C, let me fill these up, R2.1557

Now, the S, these electrons have moved and formed a bond here, so what I have is, I have formed a bond with this sulfur.1568

Now, this sulfur is attached to this carbon.1576

It is attached to that carbon.1581

Well, that carbon is now attached to this nitrogen with a double bond, and that nitrogen is attached to this carbon.1583

That is what's happening.1595

Also attached to this carbon is the...that's it.1597

Just keep track of your carbons.1608

That is all that's happening here.1611

Again, and this is N, C, C, so the R1 group is right here.1613

Again, let's keep track of our...OK.1620

We formed this as an intermediate species.1632

Now, the next step of the reaction is the following.1635

Let me do this in red again.1642

OK.1644

This bond is the bond that we are going to break right here- right between the C and the N.1645

This is one amino acid; this is the other amino acid residue- N, C, C, N, C, C.1652

So, what happens is these electrons right here, they go back down to form the carbonyl because the carbonyl is very stable, and they kick off these bonds, and they go on to grab an H+.1659

There is an H right here, by the way.1673

And therefore, this bond is actually broken.1676

What you end up with is the following.1680

I'm going to draw this out.1690

This is going to be in blue.1692

Actually, you know what, let me draw...that's fine.1695

I'll just go ahead and do that, that's fine, but I'm going to draw this molecule over here.1702

This is going to be C…nope, do it in blue.1710

We have C, we have C, we have the carbonyl is formed again.1713

We have S, we have C, we have double...oops...we have double bonded N, single bonded C.1720

We have our R1 group, and, of course, here, we have our NH and our benzene ring, plus we have N, NH.1736

It grabbed an H, so we have NH2, C, C, O-, R2, and again, keep track, N, C, C.1750

There we go.1769

We have our amino acid; we have our derivative part, and this is the one that undergoes that rearrangement to form the final PTH; but I wanted you to see this mechanism.1771

It is the nitrogen electrons that move here to form the double bonded carbon.1788

They push the double bond on the sulfur, and attacks the carbonyl.1792

The electrons move up onto the oxygen to carry a negative charge.1795

The electrons come back down to form the carbonyl, and they kick off these electrons to have it to do whatever it does; and that actually breaks this bond right there.1798

Let me go ahead and do this in black after the fact- this bond is broken.1809

There you go.1823

That is the mechanism.1824

OK.1827

Let's go on here, see what we can do.1828

Let's go ahead and do an example.1830

OK, an example.1833

OK.1840

Write out the Edman degradation for the tripeptide Ala, Tyr, Ser- alanine, tyrosyl, serine.1843

We have this tripeptide.1873

We want you to write out structurally the Edman Degradation using arrows, not mechanism arrows.1874

We just want you to show what reagents you are using, what the products are going to be for the entire Edman degradation for this thing.1880

OK.1887

Let's just jump in, and you need to be able to do this.1888

You have to be able to reproduce this - very, very important.1889

You get practice with amino acid structures; you get practice with writing out the PITC, PTC, PTH- all of that stuff.1892

Again, you do enough of this, 3 or 4, 5 of these, you'll be perfect; but you have to do them.1900

OK.1907

Let's draw it out.1910

Let's see; let's go.1913

Shall we do it in blue or black?1915

It doesn't really matter; let's do it in blue.1917

Again, do the backbone first, N, C, C...oops...N, C, C, N, C, C.1919

And again, you can use the shape structure; you can use line structure- whatever works best for you.1931

I just love seeing everything.1935

We've got H2 or H3- it doesn't really matter.1937

We have carbonyl on the second carbon, carbonyl on the second carbon, carbonyl on the second carbon.1944

Let's go ahead and put an O- there.1948

We have alanine, which is CH3.1950

Notice, I'm not putting the H on the alpha-carbon anymore.1953

We have tyrosine.1957

I probably should have picked something a lot easier, but OK, and a little less tedious to draw out, but that's OK; it's good practice.1959

I like tyrosine and serine, which is CH2OH, if I'm not mistaken.1971

OK.1978

The first step is, you are going to use PITC, phenyl isothiocyanate, under mildly basic conditions, and you are going to form the following.1979

You are going to form N, C, S, N, H, C, C.1996

Here, this is going to be CH3, and this is going to be N, C, C, N, C, C.2014

We've got N, H, C.2022

This is going to be CH2.2026

We have OH, we have our carbonyl, and we have NH, we have CH2OH, we have that, and we have that.2031

That is our first step.2040

We have formed this thing.2041

OK.2044

This is our phenylthiohydantoin.2045

So, we've formed this PTC thing.2052

Let me just go ahead and write that in red.2053

Where do I put it?2058

That is fine; I'll just put it here.2059

We have formed PTC.2060

OK.2062

Our next step, let's go ahead and actually...that's fine; I'll just do it on the next page.2063

Let's go back to blue.2069

Now, we're going to use trifluoroacetic acid.2071

Let's just write TFA- trifluoroacetic acid.2078

When I do that, I'm going to actually form a ring, and I'm going to break a bond.2082

So, let's see which bond am I going to break.2087

Well, I have that.2091

I'm looking for N, C, C, N, C, C, N - my first peptide bond.2092

That is the bond that is going to break.2096

The ring that I'm going to form is going to be made up of sulfur, 1, 2, 3, 4, 5, 5-membered rings starting with sulfur - sulfur, carbon, nitrogen, carbon, carbon.2099

That is my 5-membered ring.2114

OK.2116

Let's go ahead and form that then.2117

Let me see; do I actually do a...yes, that's not a problem.2122

Let's go ahead and form that.2127

Let me write it out over here.2129

Let me go back to blue.2134

It is going to be C, C, carbonyl, CH3; it's going to be N, C, S, and on this C is going to be the NH, and it's going to be that thing, and what we are left with is...where am I? yes... I'm left with tyrosine and serine.2136

I'm left with H3, N+, N, C, C, N, C, C.2176

I've got a carbonyl there; I’ve got a carbonyl there.2184

This is CH2, and yes it is CH2, and this is going to be my tyrosine.2190

As you can see, keeping track of all these gets really kind of confusing.2207

So, don't feel bad if you have difficulty with this because we all do.2211

OK.2216

So, I formed this thing right here, my anilinothiazolinone; and this is what I'm going to subject to H+, under aqueous conditions.2217

And again, what we want to reverse is, this and this are going to switch places.2235

I'm going to leave everything the same; I'm just going to switch that and that.2245

I'm going to write this one in back to blue.2249

I'm going to go C, C; I'm going to go N.2252

I'm going to go C; I'm going to go N that way.2259

This one is the carbonyl; this is the alanine.2264

This C has now an S, and this actually has that phenyl group attached to it.2270

So, this is my final product; this is my PTH, my phenylthiohydantoin.2279

This is the one that I'm going to identify.2285

And again, my amino acid, N, C, C, is right here.2287

That's it.2295

Now, we go ahead and we take the next step.2297

Now, we take in this molecule, so we've gone ahead and identified one, now, we are going to subject this molecule - I hope you don't mind if I change colors here - I'm going to react now, this one for the second cycle.2299

I'm going to react it with PITC under mildly alkaline conditions, and I'm going to end up forming the following molecule.2314

Yes, that's very, very important that you write all of these out, at least a couple of times.2325

C, S, N, C, C, N, C, C, N, C, C, that carbonyl goes there, that carbonyl goes there, and here we have the tyrosine R-group, and here we have the serine R-group.2330

OK.2357

We have actually formed this bond right here with the phenyl isothiocyanate.2358

Now, this is the one that we are going to subject to trifluoroacetic acid in order to form the ring and break this bond.2365

Now, we are going to break that bond, and we are going to form a ring from this molecule.2376

Let's go ahead and form that; let's see what that looks like.2382

That is going to end up looking like this.2386

It is going to be C, C, O, we have an S, we have a C, we have double bonded N, we have that.2389

On this C, we have NH, and we have that.2402

And on this C, we have our tyrosine, CH2, and OH.2408

Now, we have plus our CH2, N, C, C.2417

We have our final serine residue which is going to be CH2OH.2425

Oops, let me do it the way that I usually do which is vertically.2430

OK.2436

That one is taken cared of, and this one is going to go on into a third cycle, which I will have you do.2437

And, let me see, this is going to be our anilinothiazolinone, which we are going to subject to aqueous acid; and when we subject it to aqueous acid, we are going to rearrange.2444

And again, what we are going to rearrange is, this thing and this thing are going to switch places.2457

Everything stays the same except those that switch places.2463

So, what I end up with is C, C, N goes there, C stays, N goes here.2466

And again, this double bond changes; it becomes a single bond that goes there.2478

That is a carbonyl; this is going to be our tyrosine group.2482

OK.2489

And this is going to happen...no, sorry about that, this and this switched, so we actually have that there.2493

This one has an H, and, of course, we have an S; but we don't want these stray lines, otherwise you are going to think that they are double bonds.2502

We don't want that.2509

So, we have that.2510

That is our final PTH, and in this particular case, our amino acid residue is right there.2513

Again, N, C, C, just follow the N, C, C, and attach the rest.2523

We can identify this, and then we just subject this to the next cycle.2530

That is it; that' all you are doing.2535

Again, it is very, very important that you do at least a couple of these by writing out the reactants, the reagents, and the product.2537

That is the only way to get a full sense of what's going on, to have full command of what's going on; and I promise you, after doing a couple of these, you'll really, really feel like you understand the material, drawing it out actively.2546

That is the only way to learn this.2561

OK.2563

Thank you for joining us here at Educator.com and Biochemistry.2564

We'll see you next time, bye-bye.2566

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