Mechanistic study of proton transfer and hysteresis in catalytic antibody 16E7 by site-directed mutagenesis and homology modeling.

Antibody 16E7 catalyzes the carbon protonation of enol ether 2 to hemiacetal 3, and the carbon deprotonation of benzisoxazole 7 to phenol 8. This antibody shows an extreme case of hysteresis, requiring several hours to reach full activity. Antibody 16E7 was expressed as recombinant chimeric Fab in Escherichia coli.

A catalytic antibody against a tocopherol cyclase inhibitor.

The cyclic ammonium cation 5 and its guanidinium analogue 4 are inhibitors of tocopherol cyclase. Monoclonal antibodies were raised against protein conjugates of the haptens 1-3 and screened for catalytic reactions with alkene 8, a short chain analogue of the natural substrate phytyl-hydroquinone 6, and its enol ether analogues 10a,b. Antibody 16E7 raised against hapten 3 was found to catalyze the hydrolysis of Z enol ether 10a to form hemiacetal 12 with an apparent rate acceleration of k(cat)/k(uncat)=1400.

On the Origin of the Low-Spin Character of Cytochrome P450 in the Resting State-Investigations of Enzyme Models with Pulse EPR and ENDOR Spectroscopy.

The P450 enzyme model 1 is a high-spin system. EPR and ENDOR spectra reveal the coordination of water to the Fe center. This is the first experimental proof that coordination of water is not the single determining factor in the stabilization of the low-spin character of the cytochrome P450 resting state.