Rational design of a functional NOR in Mb to understand reaction mechanism (581.1)

Abstract

Nitric oxide reductases (NORs) catalyze the two electron reduction of nitric oxide to nitrous oxide (2NO + 2H+ + 2e‐ N2O + H2O). Nitric oxide is a fundamental cellular signaling molecule, while nitrous oxide is both a potent greenhouse gas (310x GWP of CO2) as well as a powerful ozone depleting substance. Thus, understanding this process is both chemically and environmentally significant. The active site structure in NORs has been elucidated, but the mechanism is still unknown. Due to the inherent difficultly in studying these complex enzymes, rational design of simpler protein‐based model systems is desired. To this end, a structural and functional biosynthetic model of NOR was engineered through redesign of sperm whale myoglobin (swMb), and two generations (FeBMb1 and FeBMb2) have been successfully developed. Heme replacement with isostructural Zn protoporphyrin IX into FeBMb1 has allowed us to directly probe the interaction of FeB with NO. Utilizing numerous spectroscopic techniques, the mechanism of NO reduction by these biosynthetic models has been elucidated and is shown to follow the trans mechanism. Additionally, enzymatic assays have begun which seek to determine catalytic rate, and preliminary results are comparable to some native systems.