A Solvent Engineering Approach to the Study of Biomolecular Recognition in Heme Proteins and Enzymes

Northeastern University, Boston MA

Investigators

Abstract

Mabrouk MCB 0076044 Resonance Raman, X-ray crystallographic, and computational methods will be used to study three heme proteins, myoglobin (Mb), cytochrome c (cyt c), and horseradish peroxidase (HRP), in organic media. Studies of these systems will be directed toward four specific objectives: 1) elucidating the role of bound distal pocket water and the internal H-bond network in the redox mechanism of cytochrome c and myoglobin; 2) identifying perturbations of the active site structure and function of lyophilized enzyme HRP powders and cross-linked enzyme crystals of HRP in nonaqueous media; 3) determining the mechanism whereby small-molecule excipients such as poly(ethylene glycol) and KCl enhance the catalytic activity of lyophilized enzyme powders in nonaqueous media; and 4) elucidating the heme active site structural changes in HRP in benzene that result in the change in catalytic mechanism for HRP in benzene (from oxidative electron transfer to oxo transfer). The research will be divided into three focus areas: 1) resonance Raman structural studies of horseradish peroxidase enzyme powders and cross-linked enzyme crystals in nonaqueous media; 2) x-ray crystallographic study of bound water in cyt c and Mb and the E-S complex of HRP in benzene; and 3) computational study of heme proteins in nonaqueous media. This research will provide insight at the molecular level into the phenomenon of biomolecular recognition in heme proteins and enzymes and the mechanism of nonaqueous enzymology, a powerful new method of organic synthesis in which enzymes are used as biocatalysts in solvents other than water. Such knowledge will deepen fundamental understanding of the function of these proteins in nature and expand the utility of proteins and enzymes in biotechnology and biocatalysis.