Mechanistic Study of Cytochrome P450 Enzymes A Computational Insight

Abstract

Cytochrome P450 (CYP450) enzymes are the most versatile and ubiquitous heme proteins that catalyze a plethora of chemical reactions such as C-H activation, C-H amination, epoxidation, and oxidative transformations of various exogenous and endogenous substrates into potentially less toxic and hydrophilic molecules. Due to its chemical versatility and biological importance, it is one of the most prolific topics for biochemists and bioengineers. Although in the last few decades many seminal studies have been reported on the structure and functions of CYP450s, several mechanistic aspects of CYP450 chemistry are yet to be elucidated. Therefore, the present doctoral thesis provides a comprehensive study of several such mechanistic dilemmas and various fundamental problems related to the functions of different CYP450 enzymes using highly efficient computational chemistry tools such as Molecular Dynamics (MD) simulations, Density Functional Theory (DFT), and hybrid Quantum Mechanics/Molecular Mechanics (QM/MM) methods. In our work, we first attempted to investigate the fundamental aspect of CYP450 catalysis focusing on its catalytic cycle and then we gradually progressed to study the mechanistic issue of more recent non-natural reaction catalysis. The works shown in chapters two and three are devoted to unfolding the enigmatic mechanistic issues associated with the dioxygen diffusion and protonation machinery at the catalytic cycle. Thereafter, we moved on to study a long-standing mechanistic puzzle of C-C bond cleavage reaction facilitated by a natural CYP450 enzyme i.e. CYP51 using active oxidant Cpd I. Finally, with the wave of recent trends, we reveal the mechanistic conundrum of a bioengineered CYP450 enzyme, CYP411 that catalyzes an unorthodox C-H amination reaction selectively and efficiently. In summary, the present thesis rectifies a previous consensus of the CYP450 mechanism and provides several new mechanistic insights that may be helpful to bioengineer the CYP450 enzyme for new functions.