B.A., 1961, University of Pennsylvania;
M.D., 1968, University of Pennsylvania;
D.Phil., 1976, Molecular Biophysics, Oxford University (U.K.)
Our research is directed towards the study of the structural basis of enzyme action through a detailed correlation of the electronic structure, stereochemistry, and molecular structure of enzyme-substrate complexes. In these studies we have developed methods to stabilize catalytically active intermediates of enzyme reactions with use of organic-aqueous cosolvent mixtures at sub-zero temperatures for structural characterization by magnetic resonance methods. This information provides a picture of the structural basis of interactions between the substrate and enzyme that lead to catalysis. To further understand the structural basis of enzyme action, the interactions of the substrate with active site residues structurally defined through measurement of critical interatomic distances using magnetic resonance methods are analyzed with use of computer controlled molecular graphics and correlated with results of kinetic, chemical, and mutagenic experiments.
Our investigations have been conducted through the application of electron spin resonance (ESR) and electron nuclear double resonance (ENDOR) spectroscopy. We have shown that the accuracy of structural information obtained with ENDOR is exceeded only by that of single crystal X-ray diffraction. For a variety of enzymes we have developed an approach using methods of molecular genetics for overproduction of enzymes for random and site-specific isotopic enrichment of residues. With such specially generated enzymes we can remove the background proton signals of distant residues and selectively incorporate into the active site isotopically enriched amino acids as spectroscopic marker probes.
We have also begun a combined spectroscopic and cell biological investigation into the molecular basis and target enzymes of the insulin-like activity of organic chelates of the vanadyl ion in the insulin signaling pathway.