Chymotrypsin Mechanism

Summary

In this video, Deana, a group leader for biochemistry (BCH 4024), explains the chymotrypsin mechanism, a process crucial for peptide bond hydrolysis. She begins by reviewing key terms like nucleophiles, electrophiles, and general acids and bases, which are essential for understanding the mechanism. The process involves a catalytic triad made up of asparagine, histidine, and serine residues. Deana walks through each step, illustrating how these residues work together to break down proteins, highlighting the role of histidine as a general base and acid, and serine as a nucleophile. The mechanism is presented in two phases, each with detailed explanations of the enzyme's actions and product formation.

Takeaways

• 😀 Nucleophile vs Electrophile: Nucleophiles donate electron pairs, while electrophiles accept them. In the context of protons, a general acid donates protons, and a general base accepts protons.
• 😀 Key Amino Acid Residues: Asparagine, glutamate, lysine, tyrosine, cytosine, and histidine are capable of acting as general acids or bases, while only a few act in coenzyme catalysis.
• 😀 Chymotrypsin Overview: Chymotrypsin is a pancreatic protease that performs proteolysis by hydrolyzing peptide bonds, breaking proteins into smaller peptides.
• 😀 Protease Function: Chymotrypsin breaks peptide bonds formed in proteins during a dehydration reaction, specifically targeting the C-terminal end of aromatic polypeptides.
• 😀 Active Site: Chymotrypsin's catalytic triad includes the residues Asparagine 102, Histidine 57, and Serine 195, which are key to its enzymatic function.
• 😀 General Concept Focus: The mechanism involves understanding the roles of general acids, bases, nucleophiles, and electrophiles, not just the specific steps.
• 😀 Mechanism Phases: The process is split into two phases—first, cleaving the C-terminal product, and then handling the N-terminal product of the polypeptide bond.
• 😀 Phase 1 Positioning: The polypeptide chain aligns with the enzyme, placing the aromatic group in the hydrophobic pocket and the carbonyl group in the oxyanion hole.
• 😀 Histidine as a Base: Histidine 57 acts as a general base, deprotonating Serine 195, which becomes a nucleophile to attack the carbonyl group in the peptide bond.
• 😀 Phase 2 and Water Involvement: In phase 2, water replaces the proton from Serine 195, enabling Histidine 57 to act again as a general base and facilitate nucleophilic attack on the carbonyl group to form the N-terminal product.

Q & A

What is a nucleophile, and how is it involved in the chat tripson mechanism?

A nucleophile is a species that donates an electron pair in a chemical reaction. In the chat tripson mechanism, nucleophiles such as Serine 195 act to attack the carbonyl group in the peptide bond, breaking it to form products.

What role does histidine 57 play in the chat tripson mechanism?

Histidine 57 acts as a general base in the chat tripson mechanism. It deprotonates Serine 195, making it nucleophilic, and later acts as a general acid to protonate the leaving group, facilitating the cleavage of the peptide bond.

What is the function of the catalytic triad in the enzyme chat tripson?

The catalytic triad, made up of Asparagine 102, Histidine 57, and Serine 195, forms the active site of chat tripson. These residues work together to enable the enzyme to catalyze the hydrolysis of peptide bonds during proteolysis.

What is meant by 'general acid' and 'general base' in this context?

A general acid is a molecule that donates a proton (H+), while a general base accepts a proton. In the chat tripson mechanism, histidine 57 acts as a general base to deprotonate Serine 195 and a general acid to protonate the leaving group.

How does water contribute to the mechanism in phase two?

In phase two, water acts as a source of protons for histidine 57. It is positioned in the active site and donates a proton to histidine, allowing histidine to act as a general base once again. The water molecule then nucleophilically attacks the carbonyl group of the peptide bond, breaking it.

What is the significance of the hydrophobic pocket in the enzyme’s active site?

The hydrophobic pocket in the enzyme’s active site is where the aromatic group of the substrate polypeptide chain binds. This helps position the substrate properly for catalysis, ensuring that the correct peptide bond is cleaved by the enzyme.

How does the oxyanion hole function during the mechanism?

The oxyanion hole stabilizes the negative charge that develops on the oxygen atom of the carbonyl group during the nucleophilic attack. This interaction helps to stabilize the transition state and facilitates the cleavage of the peptide bond.

What is meant by 'coenzyme catalysis' as mentioned in the transcript?

Coenzyme catalysis refers to a temporary bond formed between the enzyme and the substrate, helping to facilitate the reaction. In this case, specific residues like histidine and serine participate in transient interactions to catalyze the hydrolysis of the peptide bond.

Why is the reaction in the chat tripson mechanism considered a hydrolysis?

The reaction is a hydrolysis because it involves the breaking of a peptide bond through the addition of water. This process results in the formation of two peptide fragments and the regeneration of the enzyme.

How does the enzyme regenerate for the next cycle of catalysis?

The enzyme regenerates after each phase by exchanging protons between histidine, serine, and water. This allows the active site to reset, enabling the enzyme to catalyze additional rounds of peptide bond cleavage.