Staphylococcus aureus is a Gram-positive pathogen that is responsible for causing a variety of illness such as meningitis and septicemia. In order to mount a successful infection, S. aureus requires iron to grow; however, humans have developed a defense mechanism to limit the amount of iron available for bacterial use. Most of the body’s iron content is found in heme (protophorphyrin IX + iron) bound to hemoglobin. During an infection, S. aureus preferentially utilizes hemoglobin as an iron source. This microbe uses the iron-regulated surface determinant protein H, IsdH, to rapidly extract heme from hemoglobin. A highly conserved unit within IsdH extracts heme from hemoglobin (Hb). It is formed by the second (N2) and third (N3) near iron transporter (NEAT) domains that bind to hemoglobin and heme, respectively. The domains are connected by a helical linker domain, whose function in the extraction process is poorly understood. A recent nuclear magnetic resonance (NMR) study of the apo-form of IsdH revealed inter-domain mobility of the N2 domain compared to the rest of the protein. To determine the range of N2’s mobility and understand its affect on IsdH’s function, residues connecting the N2 domain to the linker-N3 domains were replaced with a series of glycine-serine repeats in effort to increase this interface’s flexibility. 15N-relaxation NMR experiments were performed and reveal an increase of flexibility at this interface, and a slight increase in the mobility of the Hb binding N2 domain. Using a heme extraction assay, we demonstrate that increased flexibility in the receptor reduces the overall efficiency of heme extraction from Hb as compared to the wild type IsdH receptor. Future studies will seek to define the molecular basis through which increased domain flexibility impairs receptor function.

Understanding the role of inter-domain dynamics of the Staphylococcus aureus hemoglobin receptor, IsdH, and its influence on the heme extraction mechanism

251st National Meeting (2016)

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