Structural basis for iron piracy by pathogenic Neisseria

Abstract

Neisseria are obligate human pathogens causing bacterial meningitis, septicaemia and gonorrhoea. Neisseria require iron for survival and can extract it directly from human transferrin for transport across the outer membrane. The transport system consists of TbpA, an integral outer membrane protein, and TbpB, a co-receptor attached to the cell surface; both proteins are potentially important vaccine and therapeutic targets. Two key questions driving Neisseria research are how human transferrin is specifically targeted, and how the bacteria liberate iron from transferrin at neutral pH. To address these questions, we solved crystal structures of the TbpA–transferrin complex and of the corresponding co-receptor TbpB. We characterized the TbpB–transferrin complex by small-angle X-ray scattering and the TbpA–TbpB–transferrin complex by electron microscopy. Our studies provide a rational basis for the specificity of TbpA for human transferrin, show how TbpA promotes iron release from transferrin, and elucidate how TbpB facilitates this process.

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Figure 1: Crystal structure of the TbpA–(apo)hTF complex.
Figure 2: TbpA distorts the iron coordination site in the hTF C lobe by inserting a helix from extracellular loop three.
Figure 3: SAXS analysis of the N. meningitidis TbpB–(holo)hTF complex.
Figure 4: Analysis of the TbpA–TbpB–(holo)hTF triple complex by negative stain electron microscopy.
Figure 5: Mechanism for iron import.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

Coordinates and structure factors for TbpA–(apo)hTF, TbpA–(apo)hTF C lobe, diferric hTF, apo-hTF C lobe and TbpB are deposited in the Protein Data Bank under accession codes 3V8X, 3V89, 3V83, 3SKP and 3V8U, respectively.

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Acknowledgements

N.N., N.C.E., M.O., E.B. and S.K.B. are supported by the Intramural Research Program of the NIH, National Institute of Diabetes and Digestive and Kidney Diseases. M.O. was initially funded by an EPSRC Research Committee Studentship awarded to S.K.B. and R.W.E. N.M. and A.C.S. are supported by the Intramural Research Program of the NIH, National Institute of Arthritis and Musculoskeletal and Skin Diseases. A.B.M. was supported in part by USPHS grant R01-DK21739. A.N.S. is funded by an AHA Predoctoral Fellowship (10PRE4200010). E.T. acknowledges NIH support by R01-GM086749, U54-GM087519 and P41-RR05969. All the simulations were performed using TeraGrid resources (MCA06N060). We thank the respective staffs at the Southeast Regional Collaborative Access Team (SER-CAT) and General Medicine and Cancer Institutes Collaborative Access Team (GM/CA-CAT) beamlines at the Advanced Photon Source, Argonne National Laboratory for their assistance during data collection. Use of the Advanced Photon Source was supported by the US Department of Energy, Office of Science, Office of Basic Energy Sciences, under Contract No. W-31-109-Eng-38 (SER-CAT), and by the US Department of Energy, Basic Energy Sciences, Office of Science, under contract No. DE-AC02-06CH11357 (GM/CA-CAT). Portions of this research were carried out at the Stanford Synchrotron Radiation Laboratory, a national user facility operated by Stanford University on behalf of the US Department of Energy, Office of Basic Energy Sciences. The SSRL Structural Molecular Biology Program is supported by the Department of Energy, Office of Biological and Environmental Research, and by the National Institutes of Health, National Center for Research Resources, Biomedical Technology Program.

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N.N., N.C.E., M.O. and S.K.B. expressed, purified and crystallized TbpA, TbpB and various hTFs. N.N. solved all crystal structures and the SAXS structure and analysed all data. A.B.M. and A.N.S. designed and purified apo-hTF, holo-hTF, hTF–FeN and hTF–FeC for binding experiments with TbpA and TbpB; they also expressed and purified hTF C lobe for the corresponding structure (PDB code 3SKP). P.A. and O.Z. expressed and purified hTF C lobe for the TbpA–(apo)hTF C-lobe structure (PDB code 3V89). N.M. and A.C.S. designed, conducted and analysed EM experiments. E.T. and J.G. designed, conducted and analysed molecular dynamics simulations. E.B. participated in the data collection and analysis of the SAXS data. R.W.E., A.R.G. and S.K.B. conceived and designed the original project. N.N. and S.K.B. wrote the manuscript.

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Correspondence to Susan K. Buchanan.

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Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-18 with legends, Supplementary Tables 1-4, full legends for Supplementary Movies 1-2 and additional references. (PDF 28815 kb)

Supplementary Movie 1

The movie shows the molecular dynamics simulation of the TbpA-TonB interaction (see Supplementary Information file for full legend). (MOV 19599 kb)

Supplementary Movie 2

The movie shows the iron import machinery from pathogenic Neisseria (see Supplementary Information file for full legend). (MOV 29868 kb)

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Noinaj, N., Easley, N., Oke, M. et al. Structural basis for iron piracy by pathogenic Neisseria. Nature 483, 53–58 (2012). https://doi.org/10.1038/nature10823

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