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Peroxidasin forms sulfilimine chemical bonds using hypohalous acids in tissue genesis

Nature Chemical Biology volume 8, pages 784–790 (2012)Cite this article

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Abstract

Collagen IV comprises the predominant protein network of basement membranes, a specialized extracellular matrix, which underlie epithelia and endothelia. These networks assemble through oligomerization and covalent crosslinking to endow mechanical strength and shape cell behavior through interactions with cell-surface receptors. A recently discovered sulfilimine (S=N) bond between a methionine sulfur and hydroxylysine nitrogen reinforces the collagen IV network. We demonstrate that peroxidasin, an enzyme found in basement membranes, catalyzes formation of the sulfilimine bond. Drosophila peroxidasin mutants have disorganized collagen IV networks and torn visceral muscle basement membranes, pointing to a critical role for the enzyme in tissue biogenesis. Peroxidasin generates hypohalous acids as reaction intermediates, suggesting a paradoxically anabolic role for these usually destructive oxidants. This work highlights sulfilimine bond formation as what is to our knowledge the first known physiologic function for peroxidasin, a role for hypohalous oxidants in tissue biogenesis, and a possible role for peroxidasin in inflammatory diseases.

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Figure 1: PFHR-9 cells produce a basement membrane collagen IV network with sulfilimine crosslinks.
Figure 2: A basement membrane peroxidase forms the collagen IV sulfilimine bond.
Figure 3: Peroxidasin forms hypohalous acids and sulfilimine bonds in collagen IV.
Figure 4: Hypohalous acids form collagen IV sulfilimine bonds.
Figure 5: Peroxidasin uniquely crosslinks native collagen IV networks.
Figure 6: Peroxidasin is critical for collagen IV and basement membrane integrity.

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References

  1. Hynes, R.O. The extracellular matrix: not just pretty fibrils. Science 326, 1216–1219 (2009).

    Article  CAS  Google Scholar 

  2. Yurchenco, P.D. Basement membranes: cell scaffoldings and signaling platforms. Cold Spring Harb. Perspect. Biol. 3, a004911 (2011).

    Article  Google Scholar 

  3. Khoshnoodi, J., Pedchenko, V. & Hudson, B.G. Mammalian collagen IV. Microsc. Res. Tech. 71, 357–370 (2008).

    Article  CAS  Google Scholar 

  4. Vanacore, R. et al. A sulfilimine bond identified in collagen IV. Science 325, 1230–1234 (2009).

    Article  CAS  Google Scholar 

  5. Pedchenko, V. et al. Molecular architecture of the Goodpasture autoantigen in anti-GBM nephritis. N. Engl. J. Med. 363, 343–354 (2010).

    Article  CAS  Google Scholar 

  6. Fessler, L.I. & Fessler, J.H. Identification of the carboxyl peptides of mouse procollagen IV and its implications for the assembly and structure of basement membrane procollagen. J. Biol. Chem. 257, 9804–9810 (1982).

    CAS  PubMed  Google Scholar 

  7. Nelson, R.E. et al. Peroxidasin: a novel enzyme-matrix protein of Drosophila development. EMBO J. 13, 3438–3447 (1994).

    Article  CAS  Google Scholar 

  8. Adeniyi-Jones, S.K. & Karnovsky, M.L. Oxidative decarboxylation of free and peptide-linked amino acids in phagocytizing guinea pig granulocytes. J. Clin. Invest. 68, 365–373 (1981).

    Article  CAS  Google Scholar 

  9. Nagasaka, A. & Hidaka, H. Effect of antithyroid agents 6-propyl-2-thiouracil and 1-mehtyl-2-mercaptoimidazole on human thyroid iodine peroxidase. J. Clin. Endocrinol. Metab. 43, 152–158 (1976).

    Article  CAS  Google Scholar 

  10. Alexander, N.M. Iodide peroxidase in rat thyroid and salivary glands and its inhibition by antithyroid compounds. J. Biol. Chem. 234, 1530–1533 (1959).

    CAS  PubMed  Google Scholar 

  11. Weiss, S.J., Klein, R., Slivka, A. & Wei, M. Chlorination of taurine by human neutrophils. Evidence for hypochlorous acid generation. J. Clin. Invest. 70, 598–607 (1982).

    Article  CAS  Google Scholar 

  12. Tang, S.S., Trackman, P.C. & Kagan, H.M. Reaction of aortic lysyl oxidase with β-aminopropionitrile. J. Biol. Chem. 258, 4331–4338 (1983).

    CAS  PubMed  Google Scholar 

  13. Candi, E. et al. Biochemical, structural, and transglutaminase substrate properties of human loricrin, the major epidermal cornified cell envelope protein. J. Biol. Chem. 270, 26382–26390 (1995).

    Article  CAS  Google Scholar 

  14. Ortiz de Montellano, P.R., David, S.K., Ator, M.A. & Tew, D. Mechanism-based inactivation of horseradish peroxidase by sodium azide. Formation of meso-azidoprotoporphyrin IX. Biochemistry 27, 5470–5476 (1988).

    Article  CAS  Google Scholar 

  15. Rostovtsev, V.V., Green, L.G., Fokin, V.V. & Sharpless, K.B. A stepwise huisgen cycloaddition process: copper(I)-catalyzed regioselective “ligation” of azides and terminal alkynes. Angew. Chem. Int. Edn Engl. 41, 2596–2599 (2002).

    Article  CAS  Google Scholar 

  16. Obinger, C. Chemistry and biology of human peroxidases. Arch. Biochem. Biophys. 445, 197–198 (2006).

    Article  CAS  Google Scholar 

  17. Dypbukt, J.M. et al. A sensitive and selective assay for chloramine production by myeloperoxidase. Free Radic. Biol. Med. 39, 1468–1477 (2005).

    Article  CAS  Google Scholar 

  18. Morin, X., Daneman, R., Zavortink, M. & Chia, W. A protein trap strategy to detect GFP-tagged proteins expressed from their endogenous loci in Drosophila. Proc. Natl. Acad. Sci. USA 98, 15050–15055 (2001).

    Article  CAS  Google Scholar 

  19. Gotenstein, J.R. et al. The C. elegans peroxidasin PXN-2 is essential for embryonic morphogenesis and inhibits adult axon regeneration. Development 137, 3603–3613 (2010).

    Article  CAS  Google Scholar 

  20. Gupta, M.C., Graham, P.L. & Kramer, J.M. Characterization of α1(IV) collagen mutations in Caenorhabditis elegans and the effects of α1 and α2(IV) mutations on type IV collagen distribution. J. Cell Biol. 137, 1185–1196 (1997).

    Article  CAS  Google Scholar 

  21. Cheng, G., Salerno, J.C., Cao, Z., Pagano, P.J. & Lambeth, J.D. Identification and characterization of VPO1, a new animal heme-containing peroxidase. Free Radic. Biol. Med. 45, 1682–1694 (2008).

    Article  CAS  Google Scholar 

  22. Coupry, I. et al. Ophthalmological features associated with COL4A1 mutations. Arch. Ophthalmol. 128, 483–489 (2010).

    Article  Google Scholar 

  23. Favor, J. et al. Type IV procollagen missense mutations associated with defects of the eye, vascular stability, the brain, kidney function and embryonic or postnatal viability in the mouse, Mus musculus: an extension of the Col4a1 allelic series and the identification of the first two Col4a2 mutant alleles. Genetics 175, 725–736 (2007).

    Article  CAS  Google Scholar 

  24. Gould, D.B., Marchant, J.K., Savinova, O.V., Smith, R.S. & John, S.W. Col4a1 mutation causes endoplasmic reticulum stress and genetically modifiable ocular dysgenesis. Hum. Mol. Genet. 16, 798–807 (2007).

    Article  CAS  Google Scholar 

  25. Labelle-Dumais, C. et al. COL4A1 mutations cause ocular dysgenesis, neuronal localization defects, and myopathy in mice and Walker-Warburg syndrome in humans. PLoS Genet. 7, e1002062 (2011).

    Article  CAS  Google Scholar 

  26. Van Agtmael, T. et al. Dominant mutations of Col4a1 result in basement membrane defects which lead to anterior segment dysgenesis and glomerulopathy. Hum. Mol. Genet. 14, 3161–3168 (2005).

    Article  CAS  Google Scholar 

  27. Péterfi, Z. et al. Peroxidasin is secreted and incorporated into the extracellular matrix of myofibroblasts and fibrotic kidney. Am. J. Pathol. 175, 725–735 (2009).

    Article  Google Scholar 

  28. Beal, J.L., Foster, S.B. & Ashby, M.T. Hypochlorous acid reacts with the N-terminal methionines of proteins to give dehydromethionine, a potential biomarker for neutrophil-induced oxidative stress. Biochemistry 48, 11142–11148 (2009).

    Article  CAS  Google Scholar 

  29. Peskin, A.V., Turner, R., Maghzal, G.J., Winterbourn, C.C. & Kettle, A.J. Oxidation of methionine to dehydromethionine by reactive halogen species generated by neutrophils. Biochemistry 48, 10175–10182 (2009).

    Article  CAS  Google Scholar 

  30. Armesto, X.L., Canle, M., Fernandez, M.I., Garcia, M.V. & Santaballa, J.A. First steps in the oxidation of sulfur-containing amino acids by hypohalogenation: very fast generation of intermediate sulfenyl halides and halosulfonium cations. Tetrahedron 56, 1103–1109 (2000).

    Article  CAS  Google Scholar 

  31. Lavine, T.F. The formation, resolution, and optical properties of the diastereoisomeric sulfoxides derived from L-methionine. J. Biol. Chem. 169, 477–491 (1947).

    CAS  PubMed  Google Scholar 

  32. Huwiler, M., Burgi, U. & Kohler, H. Mechanism of enzymatic and non-enzymatic tyrosine iodination. Inhibition by excess hydrogen peroxide and/or iodide. Eur. J. Biochem. 147, 469–476 (1985).

    Article  CAS  Google Scholar 

  33. Blair-Johnson, M., Fiedler, T. & Fenna, R. Human myeloperoxidase: structure of a cyanide complex and its interaction with bromide and thiocyanate substrates at 1.9 A resolution. Biochemistry 40, 13990–13997 (2001).

    Article  CAS  Google Scholar 

  34. Andrews, P.C. & Krinsky, N.I. A kinetic analysis of the interaction of human myeloperoxidase with hydrogen peroxide, chloride ions, and protons. J. Biol. Chem. 257, 13240–13245 (1982).

    CAS  PubMed  Google Scholar 

  35. Taurog, A. & Dorris, M.L. Myeloperoxidase-catalyzed iodination and coupling. Arch. Biochem. Biophys. 296, 239–246 (1992).

    Article  CAS  Google Scholar 

  36. Garver, L.S., Xi, Z. & Dimopoulos, G. Immunoglobulin superfamily members play an important role in the mosquito immune system. Dev. Comp. Immunol. 32, 519–531 (2008).

    Article  CAS  Google Scholar 

  37. Zamocky, M., Jakopitsch, C., Furtmuller, P.G., Dunand, C. & Obinger, C. The peroxidase-cyclooxygenase superfamily: reconstructed evolution of critical enzymes of the innate immune system. Proteins 72, 589–605 (2008).

    Article  CAS  Google Scholar 

  38. Brownlee, M. Biochemistry and molecular cell biology of diabetic complications. Nature 414, 813–820 (2001).

    Article  CAS  Google Scholar 

  39. Touyz, R.M. & Briones, A.M. Reactive oxygen species and vascular biology: implications in human hypertension. Hypertens. Res. 34, 5–14 (2011).

    Article  CAS  Google Scholar 

  40. Yokoyama, M. Oxidant stress and atherosclerosis. Curr. Opin. Pharmacol. 4, 110–115 (2004).

    Article  CAS  Google Scholar 

  41. Bai, Y.P. et al. Role of VPO1, a newly identified heme-containing peroxidase, in ox-LDL induced endothelial cell apoptosis. Free Radic. Biol. Med. 51, 1492–1500 (2011).

    Article  CAS  Google Scholar 

  42. Shi, R. et al. Involvement of vascular peroxidase 1 in angiotensin II–induced vascular smooth muscle cell proliferation. Cardiovasc. Res. 91, 27–36 (2011).

    Article  CAS  Google Scholar 

  43. Brandes, R.P. Vascular peroxidase 1/peroxidasin: a complex protein with a simple function? Cardiovasc. Res. 91, 1–2 (2011).

    Article  CAS  Google Scholar 

  44. Lau, D. & Baldus, S. Myeloperoxidase and its contributory role in inflammatory vascular disease. Pharmacol. Ther. 111, 16–26 (2006).

    Article  CAS  Google Scholar 

  45. Li, H. et al. Microbicidal activity of vascular peroxidase 1 in human plasma via generation of hypochlorous acid. Infect. Immunity 80, 2528–2537 (2012).

    Article  CAS  Google Scholar 

  46. Kramer, R.H., Bensch, K.G., Davison, P.M. & Karasek, M.A. Basal lamina formation by cultured microvascular endothelial cells. J. Cell Biol. 99, 692–698 (1984).

    Article  CAS  Google Scholar 

  47. Weber, S., Dölz, R., Timpl, R., Fessler, J.H. & Engel, J. Reductive cleavage and reformation of the interchain and intrachain disulfide bonds in the globular hexameric domain NC1 involved in network assembly of basement membrane collagen (type IV). Eur. J. Biochem. 175, 229–236 (1988).

    Article  CAS  Google Scholar 

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Acknowledgements

This work was supported by the US National Institutes of Health (RO1 DK18381, DK18381-38S1 and 2PO1 DK065123 to B.G.H.), the Mount Desert Island Biological Laboratory Salisbury Cove Research Fund and F.H. Epstein Fellowship (to B.G.H.), the Vanderbilt Division of Nephrology Faculty Development Fund (to R.M.V.) and a Vanderbilt Physician Scientist Development Award (to G.B.). We are grateful to K.L. Rose and W.H. McDonald of the Mass Spectrometry Resource Center at Vanderbilt University for assistance with mass spectrometry, which was supported in part by the Vanderbilt Academic Venture Capital Fund. We acknowledge A. Chisholm and A. Page-McCaw for fruitful discussions during the writing of this manuscript. We thank M. Geizst (Semmelweis University) for the human peroxidasin coding sequence and L. Cooley (Yale University) for the vikingG454 protein trap Drosophila line. P. Todd, N. Danylevych and C. Venkov provided technical assistance.

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Author notes

  1. Gautam Bhave, Christopher F Cummings and Roberto M Vanacore: These authors contributed equally to this work.

Authors and Affiliations

  1. Division of Nephrology and Hypertension, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee

    Gautam Bhave, Roberto M Vanacore, Isi A Ero-Tolliver, Mohamed Rafi, Vadim Pedchenko & Billy G Hudson

  2. Center for Matrix Biology, Vanderbilt University Medical Center, Nashville, Tennessee

    Gautam Bhave, Roberto M Vanacore, Isi A Ero-Tolliver, Mohamed Rafi, Vadim Pedchenko & Billy G Hudson

  3. Department of Biochemistry, Vanderbilt University Medical Center, Nashville, Tennessee

    Christopher F Cummings & Billy G Hudson

  4. Molecular Biology Institute, University of California–Los Angeles, Los Angeles, California

    Chino Kumagai-Cresse, Liselotte I Fessler & John H Fessler

  5. Department of Molecular, Cell, and Developmental Biology, University of California–Los Angeles, Los Angeles, California

    Chino Kumagai-Cresse, Liselotte I Fessler & John H Fessler

  6. Division of Metabolic Diseases, Center for Biomedical Sciences, Korean National Institute of Health, Cheongwon-gun, South Korea

    Jeong-Suk Kang

  7. Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee

    Billy G Hudson

  8. Vanderbilt Institute of Chemical Biology, Vanderbilt University Medical Center, Nashville, Tennessee

    Billy G Hudson

  9. Vanderbilt Ingram Cancer Center, Vanderbilt University Medical Center, Nashville, Tennessee

    Billy G Hudson

  10. Kathryn W. Davis Center for Regenerative Biology and Medicine, Mount Desert Island Biological Laboratory, Salisbury Cove, Maine

    Billy G Hudson

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  1. Gautam Bhave
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Contributions

G.B. conducted, designed and analyzed data from the PFHR-9 cell culture experiments, purified collagen IV NC1 hexamers from Drosophila and conducted western blotting experiments on Drosophila mutants. C.F.C. conducted mechanistic experiments involving hypohalous acids and peroxidasin. R.M.V. conducted MS and analysis. L.I.F. prepared Drosophila materials, and C.K.-C. performed Drosophila genetics and confocal microscopy. I.A.E.-T. performed overlay experiments involving peroxidasin and other peroxidases. M.R. isolated collagen IV NC1 hexamers and sulfilimine-crosslinked peptides for further analysis. J.-S.K. isolated human peroxidasin expressing HEK293 stable cell lines, and V.P. established the PFHR-9 cell culture system for these studies. L.I.F. generated Drosophila mutant larvae, antibodies and protein reagents. L.I.F., J.H.F. and B.G.H. designed the study and wrote the paper along with G.B. All authors discussed the results and commented on the manuscript.

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Correspondence to Billy G Hudson.

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Bhave, G., Cummings, C., Vanacore, R. et al. Peroxidasin forms sulfilimine chemical bonds using hypohalous acids in tissue genesis. Nat Chem Biol 8, 784–790 (2012). https://doi.org/10.1038/nchembio.1038

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