Chymotrypsin Mechanism

Biochemistry Exam 1

Hydrophobic pocket

1.) Substrate binds in ___

ASP, HIS, SER

Catalytic Triad

Adjacent ; cleaved ; attack

1.) When substrate binds, the side chain of the residue ___ to the peptide bond to be ___ nestles in the hydrophobic pocket on the enzyme, positioning the peptide bond for ___

Oxyanion hole

2.) The tetrahedral intermediate transition state is stabilized by the ___

Nucleophilic alkoxide ion ; tetrahedral acyl-enzyme

2.) Interaction of SER and HIS generates a strongly ___ on SER; the ion attacks the peptide carbonyl group, forming a ___

Carbonyl oxygen ; oxyanion hole

2.) Tetrahedral acyl-enzyme formation is accompanied by a short lived negative charge on the ___ of the substrate, which is stabilized by hydrogen bonding in the ___

Acylation

2.) Formation of the short lived tetrahedral acyl-enzyme intermediate

Breaks ; collapses

3.) Peptide bond ___, transition state ___, and product 1 is released

Instability ; carbonyl oxygen

3.) ___ of the negative charge on the substrate ___ leads to collapse of the tetrahedral intermediate

Amino group ; breaking

3.) Reformation of a double bond with carbon displaces the bond between carbon and the ___ of the peptide linkage, ___ the peptide bond.

Leaving group ; protonated

3.) The amino ___ is ___ by His, facilitating its displacement

Acyl-enzyme intermediate ; Product 1

3.) This forms the ___, releases ___ made from amino acid leaving group

Deprotonation

4.) Water ___ by general base, covalent chemistry

Deprotonated ; nucleophilic

4.) An incoming water molecule is ___ by general base catalysis, generating a strongly ___ hydroxide ion

Ester ; tetrahedral ; negative

4.) Attack of hydroxide on the ___ linkage of the acyl-enzyme generates a second ___ intermediate, with oxygen in the oxyanion hole again taking on a ___ charge

Transition state ; oxyanion hole

5.) Formation of second ___ stabilized by ___

Carboxylate anion ; SER

5.) Collapse of second tetrahedral intermediate forms the second product, a ___, and displaces ___

Short-lived intermediate ; de-acylation

5.) The second transition state is a ___ formed by ___

Transition state intermediate ; product 2

6.) Collapse of ___ forms ___

Tetrahedral ; carboxylate anion

6.) Collapse of the ___ intermediate forms the second product, a ___, and displaces Ser

Free enzyme (Chymotrypsin)

7.) Dissociation of the second product from the active site regenerates ____