| Summary: | Pathogenic bacteria acquire essential iron using specialized iron acquisition
systems, such as the FbpABC transport system. The periplasmic FbpA protein delivers
iron to the ABC transporter. FbpA proteins have two domains with the iron binding site
located at the domain interface. A flexible inter-domain hinge region facilitates substrate dependent conformations. In general, the closed conformations are observed for holo FbpA proteins whereas the apo proteins exhibit increased hinge motion relative to the closed conformation. Closed conformations are likely important for initiating iron translocation across the inner membrane permease. Important bacterial pathogens such as Campylobacter jejuni and Bordetella pertussis contain previously uncharacterized FbpA proteins. Using phylogenetic analyses, six FbpA classes were defined which vary in conservation of iron site ligands and utilization of a synergistic anion. Class I includes the
anion-dependent neisserial FbpA (nFbpA). This thesis characterizes the Class III FbpA
from Campylobacterjejuni (cFbpA) and the Class II FbpA from Bordetella pertussis
(bFbpA). Visible spectroscopy showed high affinity iron binding of cFbpA. X-ray
crystallography showed anion-independent iron coordination by cFbpA using a histidine
and four tyrosine residues. Confonnational analyses in solution by small angle x-ray
scattering (SAXS) showed that cFbpA undergoes limited hinge motion in solution upon substrate binding. Furthermore, an iron uptake role is supported as a cJbpA deletion
strain, constructed from C. jejuni 81-176, exhibited impaired growth under iron-limited
conditions. Characterization of bFbpA by visible spectroscopy showed high affinity iron binding with carbonate, citrate and oxalate. Distinct holo conformations compared with the apo conformation were observed for bFbpA depending on the synergistic anion. The closed conformation holo bFbpA crystal structure shows iron coordination by carbonate and three tyrosine residues. SAXS analyses also showed that oxalate and citrate treated holo bFbpA exhibit distinct conformations from apo bFbpA in solution. Furthermore,
bFbpA undergoes large hinge motion in solution similar to nFbpA. Models for iron
transport are proposed in which these bob complexes of bFbpA and cFbpA are
candidates for initiating productive interactions with the permease.
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