Question

1. The active site of an enzyme usually consists of a pocket on the enzyme surface lined with the amino acid side chains necessary to bind the substrate and catalyze its chemical transformation. Carboxypeptidase, which sequentially removes the carboxyl-terminal amino acid residues from its peptide substrates, consists of a single chain of 307 amino acids. The two essential catalytic groups in the active site are furnished by Arg¹⁴⁵ and Glu²⁷⁰ .

a. How many amino acid residues apart are these two amino acids?

b. Explain how it is that these two amino acids, so distantly separated in the sequence, can catalyze a reaction occurring in the space of a few tenth of a nanometer?

c. If only these two catalytic groups are involved in the mechanism of hydrolysis, why is it necessary for the enzyme to contain such a large number of aa residues?

2. What level of protein structure would be disrupted by the following below

a. heat

b. strong acid

c. saturated salt solution

d. organic solvents such as alcohol and chloroform

Answer 1

a.

The number of in-between amino acids = 270 - 145 - 1 = 124 aa

b.

These amino acids are separated by 124 amino acids in the primary structure of the protein.

However, when the protein folds to its native conformation, these amino acids are brought together at the active site.

c.

The amino acids found at the catalytic site take part in the reaction.

However, the surrounding amino acids play a supportive role in creating the microenvironment that is necessary for the reaction to occur.

The two individual amino acids can not perform the catalytic function on their own in isolation.

Heat disrupts hydrogen bonds and non-polar hydrophobic interactions. So, secondary and tertiary structures are disrupted by heat.

Strong acid cleaves peptide bonds = Disrupts primary structure (If the primary structure is disrupted, all the higher order structures are disrupted as well).

Salt solutions and hydrophobic solutions also disrupt higher order protein structures other than primary structure.