Semester

Spring

Date of Graduation

2008

Document Type

Dissertation

Degree Type

PhD

College

School of Medicine

Department

Physiology, Pharmacology & Neuroscience

Committee Chair

Peter M. Gannett.

Abstract

The occurrence of atypical kinetics in cytochrome P450 reactions can confound in vitro determinations of a drug's kinetic parameters. During drug development, inaccurate kinetic parameter estimates can lead to incorrect decisions about a lead compound's potential for success. It has become widely accepted that in certain CYP subfamilies more than one molecule can occupy the active site simultaneously, in some cases resulting in enhanced substrate turnover (heteroactivation). However, the specific mechanism(s) by which dual-compound binding results in heteroactivation remain unclear. It is known that orientation of the substrate in the active site, as dictated by interactions with active site residues, plays a large role in metabolic outcome. Effector compounds have been shown in vitro to alter substrate position in the active site. Here, data obtained via in silico methods including docking, molecular dynamics, semi-empirical and ab initio quantum mechanics indicate that direct interaction between effector and substrate can play a role in stabilizing the substrate in an alternative conformation conducive to oxidation. In this study a high-throughput screening computer model of heteroactivation of flurbiprofen metabolism by CYP2C9 has been developed for the purpose of elucidating key interactions between substrate, effector, and enzyme responsible for heteroactivation in this system, as well as to predict as yet unknown activators.

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