Biochemistry Enzymes I

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

Today we are going to start our discussion of enzymes.0004

I cannot even begin to overemphasize the importance of enzymes.0009

I mean enzymes are biochemistry, so a really, really good understanding of enzymes, how they work, some of the quantitative aspects, things like that- absolutely essential to everything that goes on in biology.0016

Let's just jump right on in and spend a fair amount of time with enzymes.0032

OK, now, having said that, you see, what is going to happen is we are going to introduce enzymes, talk a little bit about the other general properties, a little bit about how they work.0037

We are going to get some examples of some enzyme reactions, and we are going to talk a little bit about the quantitative aspects, some of the kinetic aspects, things like that; but this whole idea...we are not going to get too much into it.0050

We do not want to say too much at this point.0064

This is one of those topics where you do run the risk of saying too much that is unnecessary.0068

We just want to give you a good, solid grounding in enzymes, and once we actually start to talk about the metabolic pathways - glycolysis, fatty acid breakdown, amino acid breakdown, citric acid cycle, things like that - we will be revisiting the enzymes.0075

A lot of what we ultimately want to learn about enzymes will come from all of the exposure that we get when we deal with the enzymes in the individual pathways.0091

Having said that, let's see what we can do.0103

Enzymes are, again, they are everything; enzymes are proteins- very, very large proteins.0107

Enzymes are proteins.0116

The only exception are the catalytic RNAs, which we will end up discussing in a subsequent class- molecular biology.0124

OK, now, some enzymes function as is.0148

That is without additional chemical groups to help them along, to make them work.0161

Others require something called a cofactor; excuse me.0186

Let's get comfortable here.0193

Others require something called a cofactor.0197

A cofactor can be one of several things; it can be an inorganic ion, maybe like iron(2+) or maybe magnesium(2+) or manganese(2+) etc., zinc, cobalt, something like that.0212

It can also be an organic molecule, an organic or occasionally a metallo-organic molecule called a - we usually call it a coenzyme because it is an actual molecule, it is not just 1 single metal ion - coenzyme.0233

Now, some enzymes require both ions and coenzymes.0263

OK, now, when an ion or coen...you know what, let's just call it a cofactor.0278

OK, all these names that we are throwing around, I know it can tend to be a little bit confusing.0294

The names, themselves, are not altogether that important except for the occasional quizzes that you take.0299

What is important, again, is the chemistry just understanding what the function of these things is.0305

When a cofactor, either ion or coenzyme, is tightly bound or covalently attached to the enzyme, we usually call it prosthetic group.0310

OK, now, I see.0350

The entire - let's try this again - enzyme plus cofactor is called a holoenzymes.0358

Again, just a bunch of nomenclature that is not altogether that important.0384

When you have the enzyme plus its cofactor, the thing that is all ready to go, it is called a holoenzyme.0388

The enzyme part alone without its cofactor, it is called an apoenzyme or sometimes an apoprotein.0395

OK, now, most coenzymes, they derive from vitamins.0423

Some examples would be...well, let's actually take a look at some.0444

We have this one right here; this, first one, this is riboflavin.0450

OK, this is one of the vitamins that we need to ingest in order for bodies to function properly, and the enzyme that is derived from riboflavin, this part right here, you notice, is right here.0454

OK, this looks like flavin adenine dinucleotide.0470

That is the coenzyme formed with the riboflavin.0480

We ingest the riboflavin; the body converts it to FAD.0484

This is the coenzyme; this is the active part that works in concert with the particular enzyme that it is attached to.0487

Another vitamin would be pyridoxine, and pyridoxine is the vitamin that we ingest.0495

The coenzyme, itself, is something called PLP.0502

It is called pyridoxal phosphate.0506

And again, we will be running into these; we will be getting into more details later on.0510

When we speak about specific enzymes, we are going to talk about these cofactors, what they do, how they do it.0515

We are going to get into detailed mechanisms and things like that.0520

Right now, I just wanted you to see some examples.0523

Let's see a couple of other examples; we have carbonic anhydrase.0528

I just wanted you to see actually, in this case, a ribbon diagram of the actual enzyme itself, and I do not know if you can see it here, that little zinc ion.0534

That is a prosthetic group; it is a cofactor for carbonic anhydrase, happens to just be a metallic ion, and this, of course, is a close up view.0546

This is zinc ion; you see it is coordinated to a histidine, to another histidine, to another histidine, and it is also coordinated to a hydroxide ion.0555

Within the enzyme, within the folds of the enzyme itself, there is this what you might call a partial active site.0565

This particular zinc ion is coordinated to 3 histidine residues and a hydroxide- that is it, nothing particularly strange going on here.0572

OK, now, I will just write "close up" here. "close up of the carbonic anhydrase".0582

OK, now, enzymes are classified according to the reactions they catalyze.0595

That makes sense.0612

Excuse me; OK, an international system has been agreed upon, but beware that many - if not most - common names are still in regular use.0620

Most of the time, we will be referring to things with their common names.0671

We will be calling them, for example, alcohol dehydrogenase.0675

We will not be calling it alcohol:NAD oxidoreductase.0679

That is going to be the formal name; for the most part, we are going to be using common names.0683

Now, clearly, a lot of the enzymes that were discovered early on before the international agreement of systematization, those common names, I mean they are just so entrenched in the literature.0688

They are just so entrenched in our daily use that they are never going to go anywhere.0700

So, they have formal names, but it is the common names that you are going to see them as.0705

The more recently discovered enzymes, those are the ones where you are going to actually use the systematic name instead of the common name because no common name exists for them.0709

OK, common names are still in regular use.0719

Let's see here; let's go ahead and move on to this.0724

Each new enzyme or each enzyme is given a number.0729

It is called the enzyme commission number and a systematic name.0741

It is given both a number and a name.0759

Now, the easy number has 4 digits.0763

The first number specifies the class of the enzyme/enzyme classes.0774

There are 6 classes; an example would be...and then, of course, you have your subclasses underneath that.0783

Example would be, let's just take the most basic one, EC 1.1.1.1.0792

Class 1: these are enzymes, and they are called oxidoreductases.0800

They are involved in electron transfer, oxidation-reduction.0810

They are involved in electron transfer facilitating the transfer of electrons from 1 molecule to another molecule, from 1 substrate to another substrate- that is it.0815

That first number, that is what that specifies.0834

Let's go ahead and look at a systematic name, in this particular case the EC 1111.0838

Its systematic name is going to be, as we said, alcohol:NAD⁺ oxidoreductase.0844

Oxidoreductase tells me there is a transfer of electrons.0865

The electrons are transferred from the alcohol.0869

There is a colon; they are transferred to NAD⁺.0873

That is what this means, from here to here- oxidoreductase.0877

The common name is alcohol dehydrogenase.0881

This is the enzyme that is responsible for metabolizing the alcohol that you drink, so it does not poison you.0895

OK, now, we have the 1111.0903

This tells me its class, an oxidoreductase.0909

OK, this one right here, it tells me that it is acting on an alcohol group, a COH group.0916

This one right here, the third number, it tells me that the electron acceptor is either NAD⁺ or NADP⁺, and this one right here, it is specifically NAD⁺.0932

If you had 1.1.1.2, that would be alcohol dehydrogenase NADP⁺-that is it.0950

Ultimately, what is important...I mean there are all the numbers; I mean you can look all of these up.0960

It is not a problem; you do not have to know any of these.0964

It is really important to know that the first one is what specifies the class, and your teacher will probably ask you to know at least that, the 6 classes of say, you have oxidoreductase.0966

You have liaise; you have ligase- things like that.0978

That is all that is going on here; the particular reaction that this enzyme catalyzes is the following.0982

Let's see; it takes a primary alcohol.0991

The enzyme binds 2 things; it binds the alcohol, and it binds the NAD⁺.0999

It brings them in close proximity, and it allows the transfer of electrons and hydrogen from the alcohol to the NADH - I am sorry - to the NAD⁺ to produce the following.1004

What the enzyme ends up releasing...you end up turning the alcohol into an aldehyde, and then, you have also created this thing- that is it.1017

Basically, all you have done is...what this enzyme has done is it rips away that hydrogen.1031

That hydrogen, it turns this single--bonded oxygen into a carbonyl, and then, it releases something called NADH and H⁺ into solution.1035

OK, alcohol dehydrogenase, now, of course, you probably already know this.1045

If not, ACE is the standard ending for an enzyme.1050

Alcohol dehydrogenase, pyruvate dehydrogenase, something isomerase, hexokinase, ACE, ACE, ACE, ACE, that is the enzyme.1066

In this particular case, the cofactor, this enzyme does have a cofactor.1078

The cofactor happens to be that molecule, and in this particular case, it is a cofactor that is not tightly bound to the alcohol dehydrogenase.1082

It is actually free to float around, so we do not call it a prosthetic group; but we do call it coenzyme.1094

OK, now enzymes are catalysts.1102

OK, let's go to blue here.1107

Enzymes are catalysts- that is it.1111

They are just catalysts; all they do is they speed up a reaction.1119

They speed up a reaction, but they do not affect the equilibrium position of a reaction.1127

That is very important; they allow a system to come to equilibrium faster.1141

They just do not change the equilibrium position.1148

Again, there are many reactions that cannot take place.1153

For example, if I put oxygen and hydrogen in a container, well, thermodynamically, that reaction want to happen.1156

It is very thermodynamically favorable; the problem is kinetically, it is not, because it has a very high activation energy.1169

It takes a lot for it to go over that initial hump in order to become water.1175

Thermodynamics and kinetics are not the same thing; a catalyst does not affect the equilibrium.1182

It affects how fast it reaches the equilibrium- that is it.1188

That is all a catalyst does; it just speeds things up.1192

Now, under physiological conditions, under physio conditions, bioreactions, they tend not to happen.1196

In other words, they are not too thermodynamically favorable; they tend not to happen, or they happen very slowly.1215

Again, this is why enzymes evolved; they evolved to allow for life processes to function without having to wait forever.1230

It is really, kind of, amazing; many of the reactions that give rise to life should not happen, would not happen.1243

Enzymes allow them to happen- absolutely extraordinary, powerful, powerful things.1251

It is a lot more than just speeding up a reaction; it is the extent to which they actually allow reactions that are so complex to actually take place, reaction that would never happen in the body, but enzymes allow them to happen.1257

OK, now, it does this, in other words, allow the reactions to happen by providing a place, an actually physical place and a context, an environment for the reaction to take place.1272

We call this place the active site- very, very important.1308

Where a particular something happens in the enzyme is called the active site.1314

It is where the reaction takes place; now, I want you to think about this.1318

You have got just this millions, thousands, whatever, different types of molecules floating around in your body in any given space.1324

In order for a reaction to take place, well, a couple of things need to happen.1334

The most fundamental of which is that...let's say you if you have to have this molecule react with this molecule, well, it is like being in a hugely crowded city.1339

How is it that 1 molecule on 1 side of the city is supposed to run into 1 molecule in the other side of the city?1348

How the hell is that going to happen?1354

They are not going to happen randomly, or even if they do happen to meet randomly, maybe these conditions are not exactly right for the reaction to take place.1356

What an enzyme does is it brings together this person or this molecule, that molecule.1366

It puts them in the same vicinity; it holds them close together, and it provides a context for them to actually go ahead and have a reaction.1371

That is what enzymes do, bring together molecules, provide a space for them to actually do what they are supposed to do which is react, in other words, align them properly, change this, change that, and then, facilitates that reaction by making it a little bit easier for the reaction to happen by possible providing it an alternate pathway, a pathway that it would not have otherwise thought of if they just happen to run into each other randomly.1380

That is the whole idea behind an enzyme; it provides a place and a context for this to happen.1404

We call that the active site; OK, now, the molecule or molecules an enzyme binds is called the substrate.1410

You might have more than 1 substrate; it just depends on your particular point of view.1435

If you want to consider like in the previous example with the alcohol dehydrogenase, well, the primary alcohol is one of your substrates.1443

You can consider the NAD⁺; you can call it a cofactor if you want.1450

Well, it is free to float around, so it is also a substrate; it is the second substrate.1454

Again, a lot of these things happen more to do with names; names are not altogether important.1459

What is important is what is happening; in this case, it brings together the alcohol and the NAD⁺ in order for them to do what they need to do, then it sends them of on their way.1464

The alcohol leaves as aldehyde; the NAD⁺ leaves as NADH, and the enzyme goes and does it for another pair of molecules.1473

OK, the molecule or molecules an enzyme binds is called the substrate.1482

Let's see; OK, now, the active site - you guys already have a sense, I think, of what enzymes are - is often a pocket on or within the enzyme in the enzyme.1488

OK, and because an enzyme is a protein, this pocket is lined with amino acids, which bind the substrate with weak interactions.1518

And basically, you have this pocket, and along the pocket, there are these amino acids, which are part of the protein, part of the enzyme.1556

They interact with the substrate in certain ways with weak interactions - hydrogen bonding, ion dipole, hydrophobic interactions - and they, sort of, form a little bit of a cocoon, a little support for the particular substrate; and then, it allows it to do whatever it does.1563

I will be talking a little bit more specifically about binding and weak interactions and things like that but OK, amino acids which bind the substrate with weak interactions.1582

OK, often the substrate is completely sequestered from solution, is completely sequestered from the aqueous environment.1593

In other words, when the enzyme binds the substrate, it does not allow water or anything else to come in.1620

It provides a space for only that molecule and whatever else needs to be there for the reaction to take place.1630

That is what we call sequestering; it separates it from solution, so you do not have a bunch of other stuff floating around.1636

Let's go ahead and just take a look at some of these things just to get a sense of what they might look like.1643

Let's see; here, we have an image of 3-dehydroquinate synthase.1649

In this particular case, we have 3 substrates; we have 3 things that are attached.1658

Some of the substrates are cofactors; some of them are not, again, just things that are attached to it.1662

We have here, if you can see it, this thing is the zinc.1668

It is a zinc ion, and over here, we have another substrate that looks like it is going to be the NAD⁺; and over here is the other substrate, the actual substrate itself, something called carbophosphonate.1675

And again, you notice, on here, this is, sort of, a surface image of the enzyme.1690

You notice that this particular binding pocket happens to be on the surface.1698

This one, the substrate, seems to be actually inside, so it is a pocket inside of it.1702

The enzyme has, sort of, closed itself around it, so it is not necessarily on the surface; and it looks like the zinc is on the inside too.1708

Here, the second image is, sort of...it shows a close up of a substrate, which is this thing right here, and some of its interactions with the amino acid residues that make up the active site.1716

Here, you can see that here is oxygen, oxygen on here.1731

I do not know which molecule this is, but it looks like it is interacting a little bit with this thing right here.1735

It looks like maybe a carboxyl group, maybe a glutamate, something like that, and notice, this is attached to the protein.1742

So, this is the protein, the amino acid side chain; it is the amino acid side chain that is actually interacting with weak interactions with the particular substrate, and that is it.1749

In another view of it, here, we have, sort of, a surface image.1761

Here, we have a ribbon diagram; again, you can see the substrate is actually right here, and it looks like it is actually deep inside of this particular enzyme.1765

It looks like maybe the enzyme opened up; substrate came in, and it closed itself around it.1778

That often happens; we will be talking about that a little bit later too, something called induced fit- that is it.1783

that is all it is- active site enzyme, sometimes on the surface, sometimes inside the enzyme, nothing particularly strange going on.1789

OK, let's talk a little bit about how catalysts do what they do.1799

Catalysts work by lowering activation energies- that is it.1805

You know from general chemistry that that is what catalysts do- activation energies and/or providing an alternate pathway.1820

I do not know; the thing is, by providing an alternate pathway, you are lowering the activation energies.1833

Shall we say lowering the activation energy and/or providing an alternate pathway; let's just say catalyst work by lowering activation energies.1840

How is that?1847

Let's just say and providing alternate pathways for the reaction to proceed.1852

OK, in terms of an energy diagram, let's take a look here.1870

We might have that, and we might have that, something like this.1875

This is our substrate or reactant.1883

Let's just go ahead and call it a substrate from now on because we want to get used to this nomenclature, and this is going to be the product.1888

This difference right here, the difference between here and here, this is something called the activation energy.1895

This is an energy hump that the substrate has to get over before it is a position to actually fall forward and move on to product, and this right here, this is called the transition state.1902

Well, you know what, let me do this in red.1919

This right here, it has to reach the transition state, and often, this energy barrier, this activation energy, is very, very high.1923

This energy barrier is what controls the rate of the reaction.1935

The lower that energy barrier, the faster the reaction is going to go because more molecules are going to have enough energy to get over that hump.1939

This right here, this path, this is the uncatalyzed reaction.1946

OK, this difference right here, the difference in energy from here to here, this is the normal ΔG.1950

That is the thermodynamics of that.1959

Now, the catalyzed reaction might go something like this.1963

Notice, this and this have not changed; we are still here and here.1967

The ΔG has not changed; thermodynamics does not change.1970

What has changed is this; now, your activation energy is a lot lower.1973

Because the energy is a lot lower, the reaction can proceed faster.1980

That is what a catalyst does; that is all a catalyst does.1984

It is not used up in the reaction; a catalyst itself, it goes in in one form.1990

It comes out of the reaction in the same form.1996

In between, it might experience some changes, but, itself, has never changed from beginning to end of the reaction.2000

All it does is it provides a different pathway for the reaction to proceed.2005

That is all; that is how a catalyst does what it does by reducing the activation energy, the energy it takes to get over a transition state.2009

OK, now, let's go ahead and see.2019

Well, let's do 1 more energy diagram here.2025

OK, this is substrate; this is product.2032

Let's go ahead and make that here; actually, you know what, let me go ahead and...I used blue for the uncatalyzed.2035

I will go ahead and keep it as blue for the uncatalyzed, here, here, like that, then, we might get something like this.2044

Here, what we have is we have enzyme plus substrate.2067

It is going to form something called the enzyme substrate complex.2074

Once the substrate binds to the enzyme, now, we have this thing.2078

Well, actually, you know what, let me...I think I am going to do 2 hills because I need 2 hills, boom, boom, and then, we will go there.2084

How is that?2089

Here, this is our enzyme substrate, and here, we have our enzyme product; and, of course, the enzyme is released, and the product is released.2102

The enzyme and substrate come together; they form something called the enzyme substrate complex.2121

The substrate is converted, changed to product; now, we have the enzyme product complex, and then, the enzyme and product separate.2126

Product is released, and enzyme goes back to start a new cycle.2133

It can be represented like this; the substrate starts here.2137

This first hump, in order to get to this first intermediate, this intermediate is the enzyme substrate complex, and then, it goes to the next hump, enzyme product complex and then, the next hump, in order to actually release this.2141

Here, well, I will tell you what; let's go ahead and just talk about it.2160

Intermediates are short-lived viable species in a reaction pathway.2165

Now, in this case, by pathway I mean individual reaction in a...you know what, I will not say pathway because we are going to actually use pathway in a different context.2190

Intermediates are short-lived viable species, I will say, in the progress of a reaction.2198

In other words, reactions do not necessarily happen in a single step.2215

They happen in multiple steps for a given reaction.2221

Each one of those steps, has a little transition state that you have to get over, and each one of those valleys constitutes an intermediate.2226

An intermediate is something that is there; it is short-lived.2234

It does not live very long, so in the case of an enzyme to an enzyme substrate, it has to go over a small hill to become an enzyme substrate.2239

Now, that is an intermediate; now, the enzyme substrate has to go over another little bump to become the enzyme product and then, the next bump in order to actually become the product.2247

Again, we have to differentiate between intermediate and transition state.2256

They are not the same thing; a transition state is a very small fleeing thing that the particular substrate has to pass through in order to become the next thing.2261

An intermediate is a place along the way from substrate to final product, sort of, you call it an intermediate product if you will.2272

It is something that it has to pass through; it is a low-energy point, not a high-energy point.2283

Now, conversion from 1 intermediate to the next intermediate constitutes a reaction step.2289

That constitutes a reaction step.2320

The slowest of those steps is called the rate-limiting steps.2327

If a reaction takes place in 5 steps, and 1 of those reactions is actually the slowest, that slow reaction is going to control the overall rate of the reaction because you can only go as fast as your slowest step- that is it.2336

A chain is only as strong as its weakest link; the slowest of those steps is called rate-limiting.2350

It makes perfect sense, nothing strange here, completely intuitive.2357

Now, here, we are talking about individual reactions that happen in multiple steps.2363

Most of what we discussed, when we start discussing the metabolic pathway in the second half of this course.2374

We are not going to be discussing individual reactions with multiple steps.2380

What we are going to discussing are reaction pathways.2385

You have from starting substrate to final, final, final product, there are going to be different reactions.2389

It is 1 reaction with multiple steps.2397

You only going to end up having a reaction, another reaction, another reaction, another reaction.2400

We are going to be talking about reaction pathways, metabolic pathways.2406

Of those metabolic pathways, you might have one of those reactions that is going to be rate-limiting.2412

You can have reaction 1, reaction 2, reaction 3 and so on.2420

Each one of those is going to be catalyzed by an enzyme, enzyme and enzyme.2430

From here to here is one reaction.2436

It may or may not involve more than 1 step in this 1 reaction, but the whole thing taken together, it is called a pathway.2442

Let's definitely make sure we distinguish between the individual steps of a reaction, which may have 1 step or more than 1 step and a pathway which involves multiple reactions.2452

We definitely want to distinguish from that; most of the time, we are going to be concerned with pathways.2465

We are not going to be concerning ourselves with the individual steps of each reaction.2470

Sometimes, we will when we want to get into the mechanistic details of that particular enzyme.2474

More often than not, we will not; OK, I will go ahead and stop this particular lesson here.2480

In the next lesson, we will continue on, get a little bit deeper into discussing enzymes, and then, we will take it from there and move forward.2487

Thank you so much for joining us here at Educator.com; we will see you next time, bye-bye.2495