Even if you read nothing but the titles of the references you
provided, you would have found that in every case the selection is for
binding and not for substrate turnover. In rare cases enzyme-activity
may be a side-effect of this selection.
Enzyme-substrate binding specificity is one criticaly important aspect
of enzyme function/turnover. The reason enzyme efficiency is measured
by production of product rather than binding is because it has been
technically difficult to directly measure the reversible binding of
substrate to enzyme and dissociation of enzyme from product (k1 and
k2). Suitable binders can be selected to undergo further testing for
catalytic efficiency.
Don't you think after the rather massive accusations against my
scientific qualifications, you owe it the community you represent to
dig up at least one of the "most famous" references of a phage display
experiment which selected for enzymatic activity (i.e. not binding,
and not screening)...
This is the first time you are introducing the distinction between
substrate binding affinity and chemical reactivity. An important one,
which I'll return to.
What massive accusations are you talking about John? My sole
"accusation" - as I mentioned in our telephone conversation - is that
you suffer from myopia when it comes to considering alternative
solutions. Your response is that SENS should explore solutions that
are not being considered by conventional science. My rebuttal was that
it does not matter what conventional science is looking at we should
be seeking the fastest way to obtain escape velocity. SENS must be
adaptable. It has not changed in 5 years. If you are going to
I don't have issue with your qualifications. Sadly, there are many
PhD's out there that end up becoming salesmen for PCR machines and lab
reagents. In contrast, Crick discovered the structure of DNA before he
was awarded his doctorate (for an unrelated thesis).
In any case, as I have pointed out above, turnover is but one way of
characterising an enzyme. Binding specificity to its substrate is
another.
But you raise an important point, and that is the development of an
assay that can discriminate between binding affinity versus chemical
reactivity from the results of a phage display methodology to generate
fucntional diversity. For example, to take the results of phage
mediated evolution and then link to a mechanism that can report
production of product. In the studies performed in our lab we use
streptavidin magnetic beads which can bind to biotinylated cells for
cell sorting. One could presumably - I don't know if this has been
done before - use a similar mechanism to separate small groups of
phage (by biotinylating them) and then monitoring for turnover in
individual wells. A rather crude example, admittedly, but the take
come message is that it can be done and should be done rather than
sending silly people to dig up soil samples from graves.