Denaturation - Video Tutorials & Practice Problems
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1
concept
Denaturation
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Video transcript
So at this point, we've covered from primary protein structure all the way up through tertiary protein structure. Now, before we get to quaternary protein structure, there are a few topics that we need to talk about, and one of those topics is denaturation. So we'll talk about that now. So denaturation is just the process of disrupting a protein's structure and specifically, we're talking about disrupting the protein's secondary or tertiary structure just enough to cause a loss of protein function. So, a denatured protein will not be able to perform its job. And so one thing that's not affected by denaturation is the primary protein structure. So the primary protein structure or the composition and sequence of the amino acids is not affected by denaturation. Now, denaturation can result from several different factors, from radiation to changes in temperature or even changes in pH. And it can even result from the addition of reagents that affect structure, and we'll talk about some of those reagents in our next lesson video. Now, renaturation is actually the reverse process of denaturation. So this is the process where we can take a denatured protein and regain the protein's original structure and shape. And so in our example below, we're going to distinguish between denaturation and renaturation. So what we have on the left over here is a normal protein shape, and so our normal protein shape has normal biological activity. That means that it does its job denatured protein is a denatured protein shape. And so notice that it's lost a lot of its structure. So the alpha helices that were present over here and some of the beta sheets and all of these turns and stuff like that, a lot of that structure has been lost. So may mostly secondary and tertiary structure. But again, the primary protein structure is still intact, so we still have our n terminal n and our c terminal n, and the order and sequence and composition of amino acids are all still fine when a protein is denatured. But our denatured protein has now lost all of its biological activity, activity, which means that it cannot perform its function anymore. Now, if we take our denatured protein and we go through a process of renaturation, that would be the reverse process of denaturation. So that's taking our denatured protein and regaining its original shape and function. So that would be the process of going from this air, this denatured protein over to the original protein. And so, this distinguishes between denaturation and renaturation and we'll be able to get some practice in our next practice video. So I'll see you guys there.
2
Problem
Problem
Which of the following is likely not affected if the pH of a protein solution is suddenly altered to 12?
A
Amino acid composition.
B
α-helices.
C
Motifs.
D
R-group interactions.
E
β-sheets.
3
concept
Denaturation
Video duration:
5m
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Video transcript
So in our last lesson video, we talked about how protein denaturation can be caused by several different factors, including the addition of specific reagents. So in this video, we're gonna talk about 2 specific reagents that can be used to denature a protein and those are urea and beta mercaptoethanol. And so what you'll see is that urea is actually known as a chaotropic agent. And all this really means is that it only disrupts the non covalent interactions. And so, what this means is that it's going to disrupt the non covalent interactions between our groups and that's going to affect the tertiary structure and denature the protein. So it turns out that, chiotropic agents, scientists don't really fully agree on the mechanism that it really uses, but they do know that these are molecules that ultimately disrupt the hydrogen bonding network of water molecules. And we know that in biological systems, the solvent for all of our proteins is going to be water. And so if we disrupt the hydrogen bonding network of water molecules, that's ultimately going to lead to altered protein stability, and that's going to disrupt the protein stability because it's going to disrupt the non covalent interactions. And so with beta mercaptoethanol ethanol on the other hand, which can be abbreviated by the symbol betam e, this specifically cleaves or breaks the dis sulfide bonds or the disulfide bridges, which we know are covalent r group interactions. And so, the way that beta mercaptoethanol cleaves these disulfide bonds is via a redox reaction or an oxidation reduction reaction. And so, in our example below, what we're gonna do is talk about the effects of urea and beta mercaptoethanol on protein structure. And so on the left hand side of our image, what we have is the effect of urea, and then on the right, what we have is the effect of beta mercaptoethanol. And so what you'll see is that on the left, what we have is our native protein on the far left here, and this is by native protein, what we mean is it's our normal protein which has its normal structure. So you can see it has its alpha helices, it's got some beta sheets, it's got all of its secondary structures in the correct way, and it also has its disulfide bonds which are in red, and so, the disulfide bonds are all, covalent r group interactions. And so our native protein is is gonna have normal biological activity and it's gonna be performing its job the right way. Now, if we go ahead and add urea, remember that urea is a chiotropic agent that disrupts only the non covalent interactions. Well, if we disrupt the non covalent interactions, actions, we're also disrupting the R group non covalent interactions and that's gonna disrupt our tertiary structure and we know that that's going to lead to a denatured protein and that's exactly what we have over here on the right. A denatured protein, which notice, it does not have its secondary structures anymore. But what you'll see is that its disulfide bonds are still completely intact. They have not been affected. So we still have our disulfide bonds here in red and so the disulfide bonds have been retained, even though we've added this urea. Now, what's interesting is if we remove the urea, so that would be this blue arrow here, in some situations, we're able to renature the protein. And recall that renature just means to regain the native protein structure and the native biological activity. And so we'll be able to see examples of the effects of urea in our next couple of videos when we talk about the Andfinson experiment. Now, if we want to completely destroy or, denature the, protein, then we need to disrupt all of its r group interactions, not just the non covalent r group interactions, but also the covalent r group interactions, which would be the disulfide bonds. And so, we need to be able to break these disulfide bonds that are here in red, and that's exactly where beta mercaptoethanol comes into play. So notice that over here, what we have is one cysteine molecule, and we know that the cysteine is linked via this disulfide bridge, which is in red. And so, again, with the addition of beta mercaptoethanol, which is this red area at the top, we know that if we add beta mercaptoethanol, which its structure looks like this, what we can do is reduce this disulfide bridge. And when we reduce the disulfide bridge here, we actually cleave or break the disulfide bridge and what that leads is to the formation of 2 separate cysteine, molecules. And so each cysteine, we know, has a sulfhydryl group, an s h group, and so we've got our 2 s h groups and notice that they are completely separate. They're no longer linked together via the disulfide bridge. And so that is the effect of beta mercaptoethanol. Now, again, what can happen is if we remove the beta mercaptoethanol under certain, conditions, we can reoxidize this, these sulfhydryl groups here and reform this disulfide bridge. And so, we'll be able to see examples of that in the Anne Finson experiment when we get to it shortly in our next couple of videos. And so, in our next video, we're gonna be able to get a little bit of practice. So I'll see you guys in that video.
4
Problem
Problem
Which of the following is least likely to result in protein denaturation?
A
Changing pH
B
Changing the [salt].
C
Disruption of weak interactions by boiling the protein solution.
D
Exposure to chaotropic agents & detergents (such as urea & SDS).