Question

The proteolytic enzyme, papain, differs from \(\alpha\) -chymdtrypsin in having cysteine or a labile derivative thereof as part of its active site. The enzyme is deactivated by substances that form complexes with, or react with, -SH groups and the activity is restored by reactions expected to regenerate an -SH group. Work out a schematic mechanism for cleavage of a peptide chain with papain which is in agreement with the idea that, while papain may not normally contain a free \(\mathrm{SH}\) group, it reacts readily with mercuric ion to give compounds of the type RSHgX. One of the most interesting features of papain is that more than 100 of its total of 185 amino acid residues may be removed with the aid of an aminopeptidase to give a fragment with considerable enzymic activity.

Short answer

Papain, a cysteine protease, cleaves peptide bonds in proteins through a mechanism involving its cysteine residue's -SH group at the active site. The mechanism involves the -SH group attacking the carbonyl carbon of the peptide bond, forming a tetrahedral intermediate, with subsequent peptide bond breakage and -SH group regeneration. This is consistent with papain's sensitivity to substances targeting -SH groups and its activity restoration upon -SH group regeneration. Interestingly, papain can lose over 100 of its 185 amino acid residues without losing enzymatic activity, highlighting the active site's importance.

Step-by-step solution

Understand the proteolytic enzyme papain

Papain is a cysteine protease enzyme that cleaves peptide bonds in proteins. It differs from \(\alpha\)-chymotrypsin as it has cysteine or a labile derivative at its active site. The enzyme is sensitive to substances that form complexes with or react with -SH groups, leading to deactivation. The activity can be restored by reactions expected to regenerate the -SH group.

Understand the reaction of papain with mercuric ion

One important feature of papain is that it reacts readily with mercuric ion to form compounds of the type RSHgX. This indicates that the -SH group plays a crucial role in the enzyme's active site and its activity.

Propose a schematic mechanism for cleavage of a peptide chain with papain

We know that papain has a cysteine residue in its active site, and a -SH group is involved in its activity. Here is a possible mechanism for the cleavage of a peptide chain by papain: 1. The -SH group of the cysteine attacks the carbonyl carbon of the peptide bond to be cleaved, forming a tetrahedral intermediate. 2. The peptide bond breaks, and the C-N bond is cleaved, resulting in the first part of the peptide chain being released. 3. A water molecule attacks the carbonyl carbon of the tetrahedral intermediate, forming another tetrahedral intermediate. 4. The cysteine -SH group is regenerated, and the second part of the peptide chain is released. This mechanism is in agreement with the idea that papain may not normally contain a free -SH group. Its activity can be restored by regenerating the -SH group, which plays a significant role in the reaction mechanism.

Recognize the unique feature of papain

Papain has a unique feature that over 100 of its 185 amino acid residues can be removed without losing its enzymatic activity. This suggests that the majority of the enzyme's structure is not essential for its function, and the active site, which contains the cysteine residue, plays the most crucial role in the enzyme's activity.

Key concepts

Cysteine Protease

Cysteine proteases are a type of proteolytic enzyme characterized by the presence of a key cysteine residue in their active site. This cysteine residue is essential for the enzyme's catalytic activity. It typically acts as a nucleophile to attack the carbonyl carbon of a peptide bond during the cleavage process. The presence of a thiol (-SH) group in the cysteine side chain is responsible for the unique reactivity of these enzymes, which can be inhibited by agents that react with -SH groups, leading to deactivation.

Papain, as a member of this enzyme family, showcases a remarkable ability to cleave peptide bonds, reflecting its precision and efficiency. Understanding how cysteine contributes to the action of papain underpins grasping the nuanced mechanisms through which it operates.

Peptide Bond Cleavage

Peptide bond cleavage is a crucial biochemical process carried out by proteolytic enzymes such as papain. This involves the breaking of the amide bond between two amino acid residues in a protein or peptide. The process typically begins with the nucleophilic attack on the carbonyl carbon of the target peptide bond by a cysteine residue in the active site of papain, forming a transient tetrahedral intermediate.

This is followed by a series of steps that result in the cleavage of the peptide bond and the release of the individual amino acid residues. Peptide bond cleavage by papain is a finely tuned process, essential in various biological pathways, including protein digestion and turnover.

Enzyme Active Site

The enzyme active site is the region where substrate molecules bind and undergo a chemical reaction. It contains specific amino acid residues that are precisely arranged to provide an environment conducive to catalysis. The active site is both structurally and chemically complementary to the substrate, allowing for a high degree of specificity in enzyme-substrate interaction.

In papain, the active site includes a vital cysteine residue that engages in the catalytic cleavage of peptide bonds. The specificity of the active site is such that it can recognize and bind to the appropriate peptide substrate, making it a focal point in understanding enzymatic function.

Enzyme-Substrate Complex

The formation of an enzyme-substrate complex is an essential step in the enzymatic reaction. It is characterized by the temporary binding of the substrate to the enzyme's active site. During this stage, the enzyme undergoes a conformational change to better accommodate the substrate, often referred to as 'induced fit'.

This close-fit interaction is critical for catalysis, as it brings the reactive groups into close proximity and properly orients them for the reaction. For papain, the enzyme-substrate complex is where the cysteine residue reacts with the peptide bond, initiating its cleavage.

Amino Acid Residues

Amino acid residues are the building blocks that make up the three-dimensional structure of enzymes. Each residue contributes specific chemical properties that can affect the enzyme's shape, reactivity, and interaction with substrates. In the context of papain, the relevance of these residues is highlighted by its ability to function even after a significant number of amino acids have been removed.

However, the integrity of the active site, particularly the presence of the crucial cysteine residue, is imperative for its catalytic activity. This demonstrates how the arrangement and identity of amino acid residues are critical for the function of proteolytic enzymes like papain.

Proteolytic Enzyme

Proteolytic enzymes, also known as proteases, are specialized enzymes that break down proteins by cleaving peptide bonds between amino acids. They play a pivotal role in many biological processes, including protein digestion, cell signaling, and the immune response. These enzymes can be highly specific, targeting particular sequences or types of peptide bonds.

Papain is one such proteolytic enzyme that, through its cysteine-mediated mechanism of action, cleaves peptide bonds efficiently and selectively. This activity not only demonstrates the importance of proteolytic enzymes in maintaining homeostasis but also represents their potential as therapeutic tools and in industrial applications.