Date of Award

August 2019

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Department

Biological Sciences

First Advisor

Daad Saffarini

Committee Members

Mark McBride, Sonia Bardy, Sergei Kuchin, Gyaneshwar Prasad

Abstract

Metals play crucial roles in many cellular processes where they form active centers of metabolic enzymes or participate in electron transfer reactions during respiration. At high concentrations, metals can be toxic and result in the formation of reactive oxygen species and protein denaturation. Bacteria have evolved homeostasis systems to maintain intracellular concentrations of various metals and avoid their toxic effects. The aim of this project is to identify and characterize metal homeostasis systems in the metal reducer Shewanella oneidensis MR-1. This bacterium can use metals and radionuclides as electron acceptors during anaerobic respiration and is therefore a good candidate for bioremediation of metal-contaminated environments. Furthermore, this bacterium is able to maintain low internal levels of heavy metals through the use of multiple efflux pumps such as the P-type ATPase - CopA, and the HME RND efflux pump – CzcCBA. This study aims to understand the role of these efflux pumps and their regulators in metal resistance.

Shewanella oneidensis also expresses a large number of c-type cytochromes, many of which function as terminal reductases. All of these proteins contain the typical heme-binding motif CXXCH and require the Ccm proteins for maturation. SirA, the terminal sulfite reductase, also possesses an atypical heme binding site CX15CH which requires a specialized system for heme attachment. S. oneidensis MR-1 encodes two cytochrome c synthetases (CcmF and SirE) and two apocytochrome c chaperones (CcmI and SirG). In this study we show that both apocytochrome c chaperones, CcmI and SirG, are required for the maturation of SirA and they each interact with the terminal sulfite reductase independently of each other, even in the absence of other components of the cytochrome c maturation system.

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