UCLA

Staphylococcus aureus and other clinically important species of Gram-positive bacteria have acquired resistance to commonly used antibiotics. This underscores the need for new therapeutics and a greater understanding of the molecular mechanisms that enable bacterial pathogens to cause disease. Iron acquisition systems are an attractive target for new antibiotics as many microbial pathogens must acquire this metal from their human host in order to establish an infection. The majority of the body’s iron supply (~60-80%) is found in the oxygen transporter hemoglobin (Hb) in the form of heme (iron + protoporphyrin IX). Many Gram-positive pathogens have therefore evolved mechanisms to acquire heme-iron form Hb. In Chapter 1, I survey our current understanding of the heme-acquisition strategies that are employed by different species of Gram-positive bacteria, especially those found in Staphylococcus aureus and Streptococcus pyogenes as they are the focus of my research. I further concentrate on the first step in the heme acquisition process, the extraction of heme from human Hb by receptors displayed on the surface of each microbe: Shr in S. pyogenes and IsdH in S. aureus. In chapters 2 and 3, structural and biochemical studies of the N-terminal region within the Shr receptor are presented. We show that the N-terminal region in Shr contains three autonomously folded domains. Two of these domains engage Hb and are renamed Hemoglobin Interacting Domains (HIDs). We present the crystal structure of HID2 which reveals a structurally unique Hb-binding domain. In Chapter 4, the kinetics and thermodynamics of heme extraction from Hb by the S. aureus IsdH are presented. Using this data and molecular dynamics simulations, a step-by-step reaction mechanism of the heme extraction process is proposed. In Chapter 5, the NMR-derived solution structure and backbone dynamics of the C-terminal domain of PhoP are reported. PhoP is part of the PhoPR two-component system in Mycobacterium tuberculosis, the causative agent of tuberculosis. This work contributes to an understanding of the mechanism by which PhoP regulates gene expression.