Research Interest

In our lab, we focus on the structural characterization of stable terminal states and unstable intermediates involved in the enzymatic reactions of heme proteins, including mammalian and bacteria cytochromes P450 and globins. These goals are principally achieved by resonance Raman (rR) spectroscopy, with the application of a genuinely innovative combination of “nano-disc” sampling methodology required for membrane-bound proteins, resonance Raman spectroscopy of isotopically labeled samples, and cryoradiolysis (77K) with a gamma ray source.

Nanodisc Technology

Cryoradiolysis of heme proteins

The ongoing research explicitly targets:

I. Physiologically important Human Stereoidogenic P450s: Intermediates and Mechanisms. The essential goals of this work are to generate, trap and structurally characterize enzymatic intermediates encountered in the enzymatic cycles of several important Human Stereoidogenic P450s, including several judiciously engineered mutants, designed to facilitate elucidation of the key active site elements, including substrate structure, that stabilize particular active site interactions with a given intermediate and thereby dictate overall reactivity.

II. Heme sensor proteins: Heme Distortion and Signaling Mechanisms. We utilize the rR technique to study several types of heme sensor proteins. The primary goal is to detect the structural variations of the heme active site in response to varying levels of “signal” gases, including O2 and NO. These studies highlight the mechanism of signal transduction and provide a deeper understanding of how these sensor proteins transduce signals in bacteria colonization of the human host.

III. Resonance Raman studies of biotechnologically bacterial P450s. Other significant rR studies were performed on the Bacillus megaterium cytochrome P450 BM3 enzyme and its biotechnology important variants, including the native and CO-bound derivative of the enzyme in presence of the drug molecules. Our rR data clearly demonstrated that binding of omeprazole and other drugs to these BM3 mutants leads to differing degrees of low spin to high spin state conversion of the heme iron, along with key structure changes at both heme and its peripheral groups, thereby leading to altered enzyme selectivity. This project has contributed an efficient alternative to an expensive aspect of drug and biotechnology development.

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