Okay, I looked at the study. It did in fact measure enzyme kinetics, but was unable to get units for this precise technical reason:
The absence of such activity when scFv P90C2 is expressed in soluble form, suggests that the structure of this antibody is dependent on the experimental conditions, and possibly even on stabilization caused by the link with the PIII phage–protein.
This, to be clear, does not make the antibody any less catalytic. When the antibody is attached to the phage it is an enzyme, but when it separates it is not. This creates a problem in quantitating how much of the protein is antibody versus phage, so you can’t get a well scaled clearance.
- Note that the concentration of scFv clones is not measurable due to the heterogeneity of the number of scFv presented on the surface of each phage particle.
However, they were able to measure the relative clearance, which scales with Km / Vmax…
Importantly, assessment of catalytic activity was performed with the scFv fragments displayed on the surface of phage, thus preventing us from quantitatively evaluating the catalytic parameters. The initial slope was thus used to compare the relative activity of scFv displayed on phage surface. It is important to note that such a measurement, on phage‐displayed scFv, is indispensable for the validation of the selection strategy and has to be considered as a prerequisite to the soluble expression of the selected scFv fragments.
They present a table that some of the activities, and two of them show higher activity than beta-lactamase itself at 45 nm. That is absolutely evidence of catalytic activity, that was not present before selection. The fact that this is not in soluble form doesn’t change anything. The virus started out without the function, but after selection it had the enzymatic function.
I understand you don’t want to concede anything on Axe’s argument publicly, but I’m not sure why your objections were valid. These are experiments that make very clear that there is enzymatic activity. They are even able to show it is substrate specific, and that it is roughly on par with native beta lactamase.
Right. That is what the negative control is for, and it came out really good. You are suggesting they needed to do more negative controls? Is your claim that it is really easy to get beta lactamase activity that is substrate specific, as they did here?
Okay, when you get a chance to clarify your objection let me know. I do a lot of work analyzing data veery similar to this, and it looks pretty good to me. Maybe you see something I missed though. I’m happy to be corrected.