Denaturation - Online Tutor, Practice Problems & Exam Prep

Denaturation disrupts a protein's secondary and tertiary structures, leading to loss of function while preserving the primary structure, which consists of the amino acid sequence. Factors like temperature, pH, and reagents such as urea and beta mercaptoethanol can induce denaturation. Urea disrupts non-covalent interactions, while beta mercaptoethanol cleaves disulfide bonds, affecting covalent interactions. Renaturation can restore the original structure and function under certain conditions, highlighting the importance of protein conformation in biological activity.

Denaturation

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. 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 one thing that's not affected by denaturation is the primary protein structure. 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. 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. What we have on the left over here is a normal protein shape, and our normal protein shape has normal biological activity. That means that it does its job. A denatured protein is a denatured protein shape. And notice that it's lost a lot of its structure. 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 mostly secondary and tertiary structure. But again, the primary protein structure is still intact, so we still have our N-terminal and our C-terminal, and the order, 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, 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 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.

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.

Denaturation

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. In this video, we're going to talk about 2 specific reagents that can be used to denature a protein, namely urea and beta mercaptoethanol. Urea is known as a chaotropic agent, which means it disrupts the noncovalent interactions. This disruption affects the tertiary structure and denatures the protein. Although scientists do not fully agree on the mechanism chaotropic agents use, they do know that these molecules disrupt the hydrogen bonding network of water molecules, which, in biological systems, is the solvent for all proteins. Disrupting this network leads to altered protein stability by affecting noncovalent interactions.

Beta mercaptoethanol, on the other hand, specifically cleaves the disulfide bonds, which are covalent R-group interactions, through a redox reaction. Below, we discuss the effects of urea and beta mercaptoethanol on protein structure. On the left, our native protein in its normal structure includes alpha helices, beta sheets, and disulfide bonds, indicated in red. Adding urea, a chaotropic agent, disrupts noncovalent interactions, leading to a denatured protein without secondary structures but retaining intact disulfide bonds. Interestingly, removing urea may sometimes allow the protein to renature, regaining its native structure and function, as we'll see in upcoming videos on the Anfinsen experiment.

To completely denature the protein, both noncovalent and covalent R-group interactions, including disulfide bonds, must be disrupted. Beta mercaptoethanol plays a crucial role here. With its addition, a disulfide bridge is cleaved, leading to the formation of two separate cysteine molecules each with a sulfhydryl group (SH). These are no longer linked by the disulfide bridge. Under specific conditions, removing beta mercaptoethanol allows the reoxidation and reformation of this disulfide bridge. We will explore further examples of this in the Anfinsen experiment in our next few videos. So, in our next lesson, we'll get some practical experience with these concepts. See you in that video.

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).

Here’s what students ask on this topic:

What is protein denaturation and how does it affect protein function?

Protein denaturation is the process of disrupting a protein's secondary and tertiary structures, leading to a loss of its biological function. During denaturation, the protein's primary structure, which is the sequence of amino acids, remains intact. Factors such as changes in temperature, pH, and the addition of specific reagents like urea and beta mercaptoethanol can cause denaturation. When a protein is denatured, it loses its specific shape, which is crucial for its function, rendering it inactive. This loss of structure means the protein can no longer perform its biological role effectively.

What are the main factors that cause protein denaturation?

Protein denaturation can be caused by several factors, including changes in temperature, pH, and the addition of specific reagents. High temperatures can disrupt the hydrogen bonds and other interactions that maintain the protein's structure. Changes in pH can alter the charge distribution on the protein, leading to structural changes. Reagents like urea disrupt non-covalent interactions, while beta mercaptoethanol cleaves disulfide bonds, affecting covalent interactions. These factors disrupt the protein's secondary and tertiary structures, leading to a loss of function.

How do urea and beta mercaptoethanol cause protein denaturation?

Urea and beta mercaptoethanol are reagents that cause protein denaturation through different mechanisms. Urea is a chaotropic agent that disrupts non-covalent interactions, such as hydrogen bonds, within the protein. This disruption affects the protein's tertiary structure, leading to denaturation. Beta mercaptoethanol, on the other hand, cleaves disulfide bonds through a redox reaction. Disulfide bonds are covalent interactions that stabilize the protein's structure. By breaking these bonds, beta mercaptoethanol disrupts the protein's tertiary structure, leading to denaturation.

Can denatured proteins be renatured? If so, how?

Yes, denatured proteins can sometimes be renatured, which means they can regain their original structure and function. Renaturation involves reversing the denaturation process by removing the denaturing agent or restoring the original conditions. For example, if a protein is denatured by urea, removing the urea can allow the protein to refold into its native structure. Similarly, if beta mercaptoethanol is used to break disulfide bonds, removing it and allowing the protein to reoxidize can restore the disulfide bonds and the protein's original structure. However, not all proteins can be renatured successfully.

What is the difference between primary, secondary, and tertiary protein structures?

The primary structure of a protein is the linear sequence of amino acids linked by peptide bonds. The secondary structure refers to local folding patterns within the protein, such as alpha helices and beta sheets, stabilized by hydrogen bonds. The tertiary structure is the overall three-dimensional shape of the protein, formed by the folding of the secondary structures and stabilized by various interactions, including hydrogen bonds, disulfide bonds, and hydrophobic interactions. Denaturation primarily affects the secondary and tertiary structures, while the primary structure remains intact.