Question

Chymotrypsin activity was measured by the hydrolysis of p- nitrophenylacetate Answer following questions

1.1. What are the catalytic triad amino acid residues? 1.2. In the reaction mechanism, what amino acid is the residue initiating the reaction mechanism, and explain how other amino acids are involved in H-bonding and acyl bond formation with the enzyme

1.3. draw the structure of tetrahedral intermediate of the substrate,

1.4. what is oxyanion hole in the active site and its role in the catalysis

1.5. explain how an water molecular is used as another substrate

Answer 1

1.1.

catalytic triad amino acid residues

Serine195, Histidine57, Aspartate102.

(number depict sequence number)

1.2 Serine is the residue initiating the reaction mechanism.

Histidine is involved in hydrogen bond formation, phenylalanine of the substrate is involved in acyl bond formation with the enzyme.

1.3

Tetrahedral structure of intermediate

1.4 An oxyanion hole is a pocket in the active site of an enzyme that stabilizes transition state negative charge on deprotonated oxygen or alkoxide. chymotrypsin contains an oxyanion hole to stabilize the tetrahedral intermediate anion formed during proteolysis and protects the substrate's negatively charged oxygen from water molecules.

1.5 Step 4 in the figure. In this, water makes the hydrogen bond with Histidine of the enzyme and the oxygen atom of water attacks the acyl ester carbon of the substrate.