Lecture 18

Question 1

Hydrolase enzymes

Answer 1

catalyze the hydrolysis of chemical bonds

Question 2

Serine Proteases

Answer 2

Distinguished by nucleophilic serine residue at the active site

• Chymotrypsin is an endoprotease
• All have the three residue triad made from a nucleophile, a general base, and an acid (usually ser(Nu), His (gen base), and asp (acid))
• binding pocket is a key feature of the mechanism

Question 3

Endoprotease

Answer 3

Peptide cleavage occurs within the chain anywhere there is an active site

Question 4

Exoprotease

Answer 4

Cleaves amino acids one at a time from either the N or C terminus

Question 5

Features of serine protease mech: Proximity effects

Answer 5

Hydrophobic pocket and catalytic triad position both the substrate and the catalytic ser-195 residue in perfect positions to promote nucleophilic attack

Question 6

Features of serine protease mech: Nucleophilic catalysis

Answer 6

Ser-195 attacks the carbonyl groups of the substrate and the intermediate acyl ester

Question 7

Features of serine protease mech: general base catalysis

Answer 7

HIs-57 accepts a proton from Ser-195, enhancing significantly the nucleophilicity of the serine OH group

Question 8

Features of serine protease mech: Stabilization of the transition state

Answer 8

the oxyanion hole formed from the amide N-H groups of Gly-293 and Ser-105 stabilize the transition states for the formation of the two tetrahedral intermediates

Question 9

Features of serine protease mech: general acid catalysis

Answer 9

The HisH+ group of His-57 donates a proton during the collapse of both tetrahedral intermediates

Question 10

Features of serine protease mech: Electrostatic catalysis

Answer 10

Asp-152 uses its negative charge to stabilize the positive charge that accumulates in His-57 during general acid base catalysis

Question 11

Site directed mutagenesis

Answer 11

method for producing mutant proteins with specific amino acid substitutions