Abstract
Lactoferrin binding protein B (LbpB) is a bi-lobed outer membrane-bound lipoprotein that comprises part of the lactoferrin (Lf) receptor complex in Neisseria meningitidis and other Gram-negative pathogens. Recent studies have demonstrated that LbpB plays a role in protecting the bacteria from cationic antimicrobial peptides due to large regions rich in anionic residues in the C-terminal lobe. Relative to its homolog, transferrin-binding protein B (TbpB), there currently is little evidence for its role in iron acquisition and relatively little structural and biophysical information on its interaction with Lf. In this study, a combination of crosslinking and deuterium exchange coupled to mass spectrometry, information-driven computational docking, bio-layer interferometry, and site-directed mutagenesis was used to probe LbpB:hLf complexes. The formation of a 1:1 complex of iron-loaded Lf and LbpB involves an interaction between the Lf C-lobe and LbpB N-lobe, comparable to TbpB, consistent with a potential role in iron acquisition. The Lf N-lobe is also capable of binding to negatively charged regions of the LbpB C-lobe and possibly other sites such that a variety of higher order complexes are formed. Our results are consistent with LbpB serving dual roles focused primarily on iron acquisition when exposed to limited levels of iron-loaded Lf on the mucosal surface and effectively binding apo Lf when exposed to high levels at sites of inflammation.
Highlights
Neisseria meningitidis is a diplococcal, Gram-negative bacteria that lives commensally in the nasopharyngeal tract of approximately 10–20% of humans [1]
The iron-hijacking process is mediated by a set of surface receptors that are specific for transferrin and lactoferrin from the host
In this study we focused on the receptors from important human pathogens responsible for meningitis and gonorrhea that are being targeted for development of vaccines, a detailed understanding of the structure and function of these proteins is needed to aid in vaccine
Summary
Neisseria meningitidis is a diplococcal, Gram-negative bacteria that lives commensally in the nasopharyngeal tract of approximately 10–20% of humans [1]. N. meningitidis is an opportunistic pathogen that can cause serious invasive infections including meningitis and sepsis. This pathogen acquires iron–an essential cofactor required for redox reactions in biological processes–from the iron-loaded host glycoproteins, human transferrin (hTf) and human lactoferrin (hLf) using a set of specialized receptors with specific affinity for these host glycoproteins [2]. The transferrin and lactoferrin receptors from N. meningitidis are both comprised of an integral outer-membrane ‘A’ protein (TbpA, LbpA) and a bi-lobed, lipidated ‘B’ protein associated with the outer membrane (TbpB, LbpB). Crystal structures of TbpB, TbpB:hTf, and TbpA:hTf from N. meningitidis have all been determined [4, 5], providing an in-depth picture of the iron acquisition pathway (Fig 1). The N-lobe of TbpB captures the iron-loaded C-lobe of hTf and brings it to TbpA where iron is removed, transported across the outer membrane, captured by FbpA and transported into the cytoplasm through the FbpBC complex
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