Bacterial Proteins Containing Novel Iron Sites

University Of Georgia (Uga), Athens GA

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Abstract

DESCRIPTION: (provided by applicant) The overall goal of this research is to determine the roles of two types of diiron-oxo proteins, ruberythrin (Rbr) and hemerythrin (Hr) homologs in the oxidative stress response of the anaerobic, sulfate-reducing bacterium, Desulfovibrio vulgaris. A comprehensive set of in vitro and in vivo experiments is proposed in order to provide the framework for understanding the roles of these proteins in the oxidative stress response. A detailed molecular characterization, including an X-ray crystal structure, of the reactive form of Rbr is proposed. Hydrogen peroxide is implicated as the substrate for Rbr. Therefore, measurements of the kinetics of oxidation of Rbr by hydrogen peroxide using stopped-flow spectrophotometry are proposed. Characterization of the ability of Rbr to function as the terminal component of a peroxidase upon combination with one or more of the ancillary proteins is also proposed. The gene for a proposed regulator of the oxidative stress response, Fur, will be probed by deletion of the gene encoding Fur in the D. vulgaris chromosome and identification of the proteins whose expression is altered in this Fur-deletion strain. Hrs have heretofore been found only in marine invertebrates, in which it functions as an oxygen uptake and storage protein. We recently discovered Hr homologs in D. vulgaris. One of these Hr homologs forms a domain of a larger protein whose role is to sense small molecules in its environment and to initiate a chemo tactic response in D. vulgaris. We propose that this chemotaxis protein, called DcrH, senses oxygen, and causes the bacteria to swim towards lower oxygen concentrations. This anaerotactic response will be tested in a dcrH deletion strain. DcrH is the first characterized Hr homolog in any microorganism. An increasing number of Rbr and Hr homologs are being discovered in infectious anaerobic microorganisms. Therefore, these proposed studies would form the basis for understanding oxidative stress not only in D. vulgaris, but also in a diverse range of anaerobic microorganisms, several of which affect human health.