Introduction to Biochemistry - Quiz 2.4.5

I failed this one with two attempts - the answer is A.

Here are my attempts:

I initially chose C because it is fairly well studied which means we can "direct" the evolution, and the fluorescence based assay make high throughput screen easy. However, I missed the keyword "multi-step". Multi-step is not great.

Then I chose D, presumably single step is better, however, radioactivity make high throughput screen hard as well.

The official correct answer is "A" - we don't know the structure nor the catalytic sites, but it is easy to screen, it is up to the evolution to help us to identify things, I guess that make directed evolution different from semi-rational approach.

It is simply $ 4 \times 4 \times 2 = 32 $.

ISM is a greedy approach, it myopically choose the current best to move on, so it is library B.

This is a trick question - the correct answer is "Not enough information". There is no way we can identify catalytic sites based on only the secondary structure, the real 3D structure is needed.

All the mutants are close relatives except mutant D, glycine to phenylalanine is a big change.