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The structure of serum resistance-associated protein and its implications for human African trypanosomiasis

Nature Microbiology volume 3, pages 295–301 (2018)Cite this article

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Abstract

Only two trypanosome subspecies are able to cause human African trypanosomiasis. To establish an infection in human blood, they must overcome the innate immune system by resisting the toxic effects of trypanolytic factor 1 and trypanolytic factor 2 (refs. 1,2). These lipoprotein complexes contain an active, pore-forming component, apolipoprotein L1 (ApoL1), that causes trypanosome cell death3. One of the two human-infective subspecies, Trypanosoma brucei rhodesiense, differs from non-infective trypanosomes solely by the presence of the serum resistance-associated protein, which binds directly to ApoL1 and blocks its pore-forming capacity3,4,5. Since this interaction is the single critical event that renders T. b. rhodesiense human- infective, detailed structural information that allows identification of binding determinants is crucial to understand immune escape by the parasite. Here, we present the structure of serum resistance-associated protein and reveal the adaptations that occurred as it diverged from other trypanosome surface molecules to neutralize ApoL1. We also present our mapping of residues important for ApoL1 binding, giving molecular insight into this interaction at the heart of human sleeping sickness.

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Fig. 1: The structure of SRA.
Fig. 2: Structural comparison of SRA with known trypanosome surface proteins.
Fig. 3: Mapping of the ApoL1 binding site of SRA by HDX-MS.
Fig. 4: Mutational analysis of SRA binding to ApoL1.

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References

  1. Rifkin, M. R. Identification of the trypanocidal factor in normal human serum: high density lipoprotein. Proc. Natl Acad. Sci. USA 75, 3450–3454 (1978).

    Article  CAS  Google Scholar 

  2. Tomlinson, S. et al. High-density-lipoprotein-independent killing of Trypanosoma brucei by human serum. Mol. Biochem. Parasitol. 70, 131–138 (1995).

    Article  CAS  Google Scholar 

  3. Vanhamme, L. et al. Apolipoprotein L-I is the trypanosome lytic factor of human serum. Nature 422, 83–87 (2003).

    Article  CAS  Google Scholar 

  4. Xong, H. V. et al. A VSG expression site-associated gene confers resistance to human serum in Trypanosoma rhodesiense. Cell 95, 839–846 (1998).

    Article  CAS  Google Scholar 

  5. Lecordier, L. et al. C-terminal mutants of apolipoprotein L-I efficiently kill both Trypanosoma brucei brucei and Trypanosoma brucei rhodesiense. PLoS Pathog. 5, e1000685 (2009).

    Article  Google Scholar 

  6. Greene, A. S. & Hajduk, S. L. Trypanosome lytic factor-1 initiates oxidation-stimulated osmotic lysis of Trypanosoma brucei brucei. J. Biol. Chem. 291, 3063–3075 (2016).

    Article  CAS  Google Scholar 

  7. Molina-Portela, M. P., Samanovic, M. & Raper, J. Distinct roles of apolipoprotein components within the trypanosome lytic factor complex revealed in a novel transgenic mouse model. J. Exp. Med. 205, 1721–1728 (2008).

    Article  CAS  Google Scholar 

  8. Thomson, R. & Finkelstein, A. Human trypanolytic factor APOL1 forms pH-gated cation-selective channels in planar lipid bilayers: relevance to trypanosome lysis. Proc. Natl Acad. Sci. USA 112, 2894–2899 (2015).

    Article  CAS  Google Scholar 

  9. Molina-Portela Mdel, P., Lugli, E. B., Recio-Pinto, E. & Raper, J. Trypanosome lytic factor, a subclass of high-density lipoprotein, forms cation-selective pores in membranes. Mol. Biochem. Parasitol. 144, 218–226 (2005).

    Article  Google Scholar 

  10. Oli, M. W., Cotlin, L. F., Shiflett, A. M. & Hajduk, S. L. Serum resistance-associated protein blocks lysosomal targeting of trypanosome lytic factor in Trypanosoma brucei. Eukaryot. Cell 5, 132–139 (2006).

    Article  CAS  Google Scholar 

  11. Hager, K. M. et al. Endocytosis of a cytotoxic human high density lipoprotein results in disruption of acidic intracellular vesicles and subsequent killing of African trypanosomes. J. Cell Biol. 126, 155–167 (1994).

    Article  CAS  Google Scholar 

  12. Perez-Morga, D. et al. Apolipoprotein L-I promotes trypanosome lysis by forming pores in lysosomal membranes. Science 309, 469–472 (2005).

    Article  CAS  Google Scholar 

  13. Vanwalleghem, G. et al. Coupling of lysosomal and mitochondrial membrane permeabilization in trypanolysis by APOL1. Nat. Commun. 6, 8078 (2015).

    Article  CAS  Google Scholar 

  14. Rifkin, M. R. Trypanosoma brucei: biochemical and morphological studies of cytotoxicity caused by normal human serum. Exp. Parasitol. 58, 81–93 (1984).

    Article  CAS  Google Scholar 

  15. Campillo, N. & Carrington, M. The origin of the serum resistance associated (SRA) gene and a model of the structure of the SRA polypeptide from Trypanosoma brucei rhodesiense. Mol. Biochem Parasitol. 127, 79–84 (2003).

    Article  CAS  Google Scholar 

  16. Stephens, N. A. & Hajduk, S. L. Endosomal localization of the serum resistance-associated protein in African trypanosomes confers human infectivity. Eukaryot. Cell 10, 1023–1033 (2011).

    Article  CAS  Google Scholar 

  17. Bart, J. M. et al. Localization of serum resistance-associated protein in Trypanosoma brucei rhodesiense and transgenic Trypanosoma brucei brucei. Cell Microbiol. 17, 1523–1535 (2015).

    Article  CAS  Google Scholar 

  18. Pays, E., Vanhollebeke, B., Uzureau, P., Lecordier, L. & Perez-Morga, D. The molecular arms race between African trypanosomes and humans. Nat. Rev. Microbiol. 12, 575–584 (2014).

    Article  CAS  Google Scholar 

  19. Higgins, M. K. & Carrington, M. Sequence variation and structural conservation allows development of novel function and immune evasion in parasite surface protein families. Protein Sci. 23, 354–265 (2014).

    Article  CAS  Google Scholar 

  20. Stodkilde, K., Torvund-Jensen, M., Moestrup, S. K. & Andersen, C. B. Structural basis for trypanosomal haem acquisition and susceptibility to the host innate immune system. Nat. Commun. 5, 5487 (2014).

    Article  Google Scholar 

  21. Higgins, M. K. et al. Structure of the trypanosome haptoglobin-hemoglobin receptor and implications for nutrient uptake and innate immunity. Proc. Natl Acad. Sci. USA 110, 1905–1910 (2013).

    Article  CAS  Google Scholar 

  22. Lane-Serff, H., MacGregor, P., Lowe, E. D., Carrington, M. & Higgins, M. K. Structural basis for ligand and innate immunity factor uptake by the trypanosome haptoglobin-haemoglobin receptor. eL ife 3, e05553 (2014).

    Article  Google Scholar 

  23. Loveless, B. C. et al. Structural characterization and epitope mapping of the glutamic acid/alanine-rich protein from Trypanosoma congolense: defining assembly on the parasite cell surface. J. Biol. Chem. 286, 20658–20665 (2011).

    Article  CAS  Google Scholar 

  24. Metcalf, P., Blum, M., Freymann, D., Turner, M. & Wiley, D. C. Two variant surface glycoproteins of Trypanosoma brucei of different sequence classes have similar 6 A resolution X-ray structures. Nature 325, 84–86 (1987).

    Article  CAS  Google Scholar 

  25. Thomson, R. et al. Evolution of the primate trypanolytic factor APOL1. Proc. Natl Acad. Sci. USA 111, 2130–2139 (2014).

    Article  CAS  Google Scholar 

  26. Li, M. Z. & Elledge, S. J. Harnessing homologous recombination in vitro to generate recombinant DNA via SLIC. Nat. Methods 4, 251–256 (2007).

    Article  CAS  Google Scholar 

  27. Wang, Z. D. et al. Universal PCR amplification of mouse immunoglobulin gene variable regions: the design of degenerate primers and an assessment of the effect of DNA polymerase 3ʹ to 5ʹ exonuclease activity. J. Immunol. Methods 233, 167–177 (2000).

    Article  CAS  Google Scholar 

  28. Fields, C. et al. Creation of recombinant antigen-binding molecules derived from hybridomas secreting specific antibodies. Nat. Protoc. 8, 1125–1148 (2013).

    Article  Google Scholar 

  29. Walter, T. S. et al. Lysine methylation as a routine rescue strategy for protein crystallization. Structure 14, 1617–1622 (2006).

    Article  CAS  Google Scholar 

  30. Kabsch, W. Xds. Acta Crystallogr. D. Biol. Crystallogr. 66, 125–132 (2010).

    Article  CAS  Google Scholar 

  31. McCoy, A. J. et al. Phaser crystallographic software. J. Appl. Crystallogr. 40, 658–674 (2007).

    Article  CAS  Google Scholar 

  32. Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D. 66, 486–501 (2010).

    Article  CAS  Google Scholar 

  33. Adams, P. D. et al. PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr. D. 66, 213–221 (2010).

    Article  CAS  Google Scholar 

  34. Bricogne, G. et al. BUSTER v.2.10.1. (Global Phasing Ltd., 2016).

  35. Winn, M. D. et al. Overview of the CCP4 suite and current developments. Acta Crystallogr. Sect. D. 67, 235–242 (2011).

    Article  CAS  Google Scholar 

  36. Murshudov, G. N., Vagin, A. A. & Dodson, E. J. Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. D. 53, 240–55 (1997).

    Article  CAS  Google Scholar 

  37. Konarev, P. V., Volkov, V. V., Sokolova, A. V., Koch, M. H. J. & Svergun, D. I. PRIMUS: a Windows PC-based system for small-angle scattering data analysis. J. Appl. Crystallogr. 36, 1277–1282 (2003).

    Article  CAS  Google Scholar 

  38. Rambo, R. P. & Tainer, J. A. Accurate assessment of mass, models and resolution by small-angle scattering. Nature 496, 477–481 (2013).

    Article  CAS  Google Scholar 

  39. Franke, D. & Svergun, D. I. DAMMIF, a program for rapid ab-initio shape determination in small-angle scattering. J. Appl. Crystallogr.  42, 342–346 (2009).

    Article  CAS  Google Scholar 

  40. Pettersen, E. F. et al. UCSF chimera – a visualization system for exploratory research and analysis. J. Comput. Chem. 25, 1605–1612 (2004).

    Article  CAS  Google Scholar 

  41. Podobnik, M. et al. Crystal structure of an invertebrate cytolysin pore reveals unique properties and mechanism of assembly. Nat. Commun. 7, 11598 (2016).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by Wellcome Trust grant 093008/Z/10/Z and MRC grant MR/L008246/1. M.K.H. is a Wellcome Trust Investigator. We thank K. Gull and S. Dean for assistance with monoclonal antibody generation. We also thank the beamline scientists at Diamond Light Source beamline I03, BioSAXS P12 beamline EMBL Hamburg and D. Staunton for assistance with biophysics equipment. S.M. and C.V.R. acknowledge with thanks funding from a MRC programme grant (MR/N020413/1) and a Wellcome Trust Instrument Grant (WT 104923/Z/14/Z) for the HDX platform.

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Authors and Affiliations

  1. Department of Biochemistry, University of Oxford, Oxford, UK

    Sebastian Zoll, Harriet Lane-Serff, Jonathan Schneider & Matthew K. Higgins

  2. Physical and Theoretical Chemistry Laboratory, University of Oxford, Oxford, UK

    Shahid Mehmood & Carol V. Robinson

  3. Department of Biochemistry, University of Cambridge, Cambridge, UK

    Mark Carrington

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Contributions

M.K.H. and S.Z. conceived and designed experiments. M.C. conducted trypanosome killing experiments. H.L.S. and S.Z. established protein purification strategies and purified proteins. J.S. cloned and purified mutant proteins. S.M. and S.Z. carried out the HDX experiments. S.M. and C.V.R. analysed and evaluated the HDX data. S.Z. collected all X-ray diffraction data (SAXS and crystallography), solved, refined and analysed the structures. S.Z. carried out the microscale thermophoresis experiments. M.K.H. and S.Z. wrote the paper with input from co-authors.

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Correspondence to Matthew K. Higgins.

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Supplementary Table 1, Supplementary Figures 1–13, Supplementary Tables 1–4.

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Zoll, S., Lane-Serff, H., Mehmood, S. et al. The structure of serum resistance-associated protein and its implications for human African trypanosomiasis. Nat Microbiol 3, 295–301 (2018). https://doi.org/10.1038/s41564-017-0085-3

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