Enzyme kinetics after site$-$specific mutagenesis

Cyclophosphamide $($CPA$)$ is an anticancer prodrug that requires activation in the liver by

cytochrome P$4502$B enzymes for production of cytotoxic metabolites. Site$-$specific

mutagenesis is used to alter the amino acid sequence of P$4502$B$1$ in an attempt to improve

the kinetics of CPA activation. The rate of reaction of CPA is studied using rat P$4502$B$1$

and a site$-$specific variant of $P4502B1$ produced using Escherichia coli. The results are as

follows.

$($a$)$ Determine the kinetic constants for the rat and site$-$specific variant enzymes.

$($b$)$ When CPA is administered to cancer patients, the peak plasma concentration of

the drug is relatively low at around $100$ to $200M.$ Are the kinetic properties of the

variant P$4502$B$1$ better than those of rat P$4502$B$1$ for CPA activation in this situation?

Explain your answer.

$($c$)$ Did manipulation of the enzyme using site$-$specific mutagenesis improve the catalytic

efficiency for CPA activation?

Hints:

Please use $\frac{1}{v}$ vs $\frac{1}{?}$ s plot to determine the kinetic constants

For $($a$),V_m$ and $K_m$ are the kinetic constants we are looking for.

For $($b$),$ lower Km means better performance at low substrate concentration situation.

For $($c$),$ catalytic efficiency is defined as $\frac{k_{\text{c}\text{a}\text{t}}}{K}m$