Mechanism of Substrate Transformations by the Complex Iron Sulfur Molybdoenzyme (CISM) Family

Indiana University, Bloomington IN

Investigators

Abstract

With this award, the Chemistry of Life Processes Program in the Chemistry Division is funding Dr. Partha Basu of Indiana University–Purdue University Indianapolis to investigate how the Complex Iron-Sulfur Molybdoenzyme (CISM) family of respiratory enzymes catalyze the reduction of a wide variety of chemicals. Respiration is an essential metabolic process that generates energy in all living systems. Many microorganisms utilize specialized enzymes to carry out anaerobic respiration where substrates such as nitrate, perchlorate, and dimethyl sulfoxide are reduced. These metabolic processes contribute to ecological diversity. Biochemical, spectroscopic, computational, and kinetic methods are used to systematically study how the structure of the active site of these CISM enzymes allows them to display substrate selectivity. This project provides graduate students and postdoctoral fellows with specialized training in enzymology, spectroscopy, and computation. In addition, the research is integrated into an outreach program that introduces high school students to the study of enzymes and attracts first-year students who are not pursuing a science degree to science. A strong emphasis of the program is to use research to convey to students from groups underrepresented in STEM careers an appreciation for science. CISM enzymes are ubiquitous and play essential physiological roles in transforming different oxyanions. This research project uses periplasmic nitrate reductase as a model enzyme and develops several mutational variants of this enzyme where the coordination environment of molybdenum is systematically modified. The kinetic analyses of the enzyme variants help develop a quantitative picture for substrate selectivity. In addition, several residues in the catalytic pocket are identified as being involved in proton shuttling and substrate binding. Mutations of these residues provide valuable information about the mechanism of the reaction. The kinetic data are placed in the context of structural and spectroscopic properties of the enzyme. The proposed research provides new detailed insights into the substrate transformations catalyzed by members of the CISM family. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.