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. 1997 Apr 15;323(Pt 2):393–399. doi: 10.1042/bj3230393

Structure of a truncated human surfactant protein D is less effective in agglutinating bacteria than the native structure and fails to inhibit haemagglutination by influenza A virus.

S Eda 1, Y Suzuki 1, T Kawai 1, K Ohtani 1, T Kase 1, Y Fujinaga 1, T Sakamoto 1, T Kurimura 1, N Wakamiya 1
PMCID: PMC1218332  PMID: 9163329

Abstract

Surfactant protein D (SP-D) is a lung-specific protein that is synthesized and secreted by lung epithelial cells and is believed to play an important role in lung host defence. This protein belongs to the C-type lectin family, which is characterized by an N-terminal cysteine-rich domain, a collagen-like domain, a neck domain and a carbohydrate recognition domain (CRD). To elucidate the biological actions of this animal lectin against such pathogens as micro-organisms, the biological activities of a recombinant partial SP-D lacking a collagen-like domain were examined. A recombinant human SP-D, consisting of a short collagen region (two repeats of Gly-Xaa-Yaa amino acid sequences), the neck domain and the CRD, was expressed in Escherichia coli. The recombinant SP-D was purified on a nickel column and then on a maltose-agarose column. This protein can form a trimeric structure owing to the neck domain and exhibits sugar-binding activity and specificity similar to those of native human SP-D. The recombinant SP-D caused dose-dependent and calcium-dependent agglutination of E. coli Y1088. The agglutination titre (the concentration required to achieve a 50% decrease in light transmission by agglutination) of recombinant SP-D was approx. 6-fold that of native SP-D. As for conglutination, the recombinant trimeric conglutinin required 8-16-fold higher concentrations than the native counterpart. In haemagglutination inhibition (HI) of influenza A virus, although native and recombinant conglutinin showed similar levels of HI activity, the recombinant SP-D was unable to inhibit haemagglutination, even at a concentration approx. 120-fold that of the native SP-D. The lectin precipitation and lectin blot assays showed that the truncated SP-D could bind to influenza A virus as well as native SP-D did. These results indicate that the agglutination activity of trimeric collectins can be largely retained, and furthermore that the oligomeric structure with several hands at opposite sites can enhance agglutination activity. The difference in HI activity against influenza A virus between native and recombinant SP-D suggests that SP-D uses a different mechanism from that of conglutinin to inhibit viral haemagglutination.

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Selected References

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  1. Anders E. M., Hartley C. A., Jackson D. C. Bovine and mouse serum beta inhibitors of influenza A viruses are mannose-binding lectins. Proc Natl Acad Sci U S A. 1990 Jun;87(12):4485–4489. doi: 10.1073/pnas.87.12.4485. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Brown-Augsburger P., Hartshorn K., Chang D., Rust K., Fliszar C., Welgus H. G., Crouch E. C. Site-directed mutagenesis of Cys-15 and Cys-20 of pulmonary surfactant protein D. Expression of a trimeric protein with altered anti-viral properties. J Biol Chem. 1996 Jun 7;271(23):13724–13730. doi: 10.1074/jbc.271.23.13724. [DOI] [PubMed] [Google Scholar]
  3. Clements J. A. Functions of the alveolar lining. Am Rev Respir Dis. 1977 Jun;115(6 Pt 2):67–71. doi: 10.1164/arrd.1977.115.S.67. [DOI] [PubMed] [Google Scholar]
  4. Drickamer K. Two distinct classes of carbohydrate-recognition domains in animal lectins. J Biol Chem. 1988 Jul 15;263(20):9557–9560. [PubMed] [Google Scholar]
  5. Eda S., Suzuki Y., Kase T., Kawai T., Ohtani K., Sakamoto T., Kurimura T., Wakamiya N. Recombinant bovine conglutinin, lacking the N-terminal and collagenous domains, has less conglutination activity but is able to inhibit haemagglutination by influenza A virus. Biochem J. 1996 May 15;316(Pt 1):43–48. doi: 10.1042/bj3160043. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hartshorn K. L., Crouch E. C., White M. R., Eggleton P., Tauber A. I., Chang D., Sastry K. Evidence for a protective role of pulmonary surfactant protein D (SP-D) against influenza A viruses. J Clin Invest. 1994 Jul;94(1):311–319. doi: 10.1172/JCI117323. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hartshorn K. L., Reid K. B., White M. R., Jensenius J. C., Morris S. M., Tauber A. I., Crouch E. Neutrophil deactivation by influenza A viruses: mechanisms of protection after viral opsonization with collectins and hemagglutination-inhibiting antibodies. Blood. 1996 Apr 15;87(8):3450–3461. [PubMed] [Google Scholar]
  8. Holmskov U., Teisner B., Willis A. C., Reid K. B., Jensenius J. C. Purification and characterization of a bovine serum lectin (CL-43) with structural homology to conglutinin and SP-D and carbohydrate specificity similar to mannan-binding protein. J Biol Chem. 1993 May 15;268(14):10120–10125. [PubMed] [Google Scholar]
  9. Hoppe H. J., Reid K. B. Collectins--soluble proteins containing collagenous regions and lectin domains--and their roles in innate immunity. Protein Sci. 1994 Aug;3(8):1143–1158. doi: 10.1002/pro.5560030801. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Khorana H. G. Rhodopsin, photoreceptor of the rod cell. An emerging pattern for structure and function. J Biol Chem. 1992 Jan 5;267(1):1–4. [PubMed] [Google Scholar]
  11. Kishore U., Wang J. Y., Hoppe H. J., Reid K. B. The alpha-helical neck region of human lung surfactant protein D is essential for the binding of the carbohydrate recognition domains to lipopolysaccharides and phospholipids. Biochem J. 1996 Sep 1;318(Pt 2):505–511. doi: 10.1042/bj3180505. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kozutsumi Y., Kawasaki T., Yamashina I. Isolation and characterization of a mannan-binding protein from rabbit serum. Biochem Biophys Res Commun. 1980 Jul 31;95(2):658–664. doi: 10.1016/0006-291x(80)90836-0. [DOI] [PubMed] [Google Scholar]
  13. Kuan S. F., Rust K., Crouch E. Interactions of surfactant protein D with bacterial lipopolysaccharides. Surfactant protein D is an Escherichia coli-binding protein in bronchoalveolar lavage. J Clin Invest. 1992 Jul;90(1):97–106. doi: 10.1172/JCI115861. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kuhlman M., Joiner K., Ezekowitz R. A. The human mannose-binding protein functions as an opsonin. J Exp Med. 1989 May 1;169(5):1733–1745. doi: 10.1084/jem.169.5.1733. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lee Y. M., Leiby K. R., Allar J., Paris K., Lerch B., Okarma T. B. Primary structure of bovine conglutinin, a member of the C-type animal lectin family. J Biol Chem. 1991 Feb 15;266(5):2715–2723. [PubMed] [Google Scholar]
  16. Lim B. L., Wang J. Y., Holmskov U., Hoppe H. J., Reid K. B. Expression of the carbohydrate recognition domain of lung surfactant protein D and demonstration of its binding to lipopolysaccharides of gram-negative bacteria. Biochem Biophys Res Commun. 1994 Aug 15;202(3):1674–1680. doi: 10.1006/bbrc.1994.2127. [DOI] [PubMed] [Google Scholar]
  17. Lu J. H., Thiel S., Wiedemann H., Timpl R., Reid K. B. Binding of the pentamer/hexamer forms of mannan-binding protein to zymosan activates the proenzyme C1r2C1s2 complex, of the classical pathway of complement, without involvement of C1q. J Immunol. 1990 Mar 15;144(6):2287–2294. [PubMed] [Google Scholar]
  18. Lu J., Willis A. C., Reid K. B. Purification, characterization and cDNA cloning of human lung surfactant protein D. Biochem J. 1992 Jun 15;284(Pt 3):795–802. doi: 10.1042/bj2840795. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Malhotra R., Haurum J. S., Thiel S., Sim R. B. Binding of human collectins (SP-A and MBP) to influenza virus. Biochem J. 1994 Dec 1;304(Pt 2):455–461. doi: 10.1042/bj3040455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Ogasawara Y., Voelker D. R. The role of the amino-terminal domain and the collagenous region in the structure and the function of rat surfactant protein D. J Biol Chem. 1995 Aug 11;270(32):19052–19058. doi: 10.1074/jbc.270.32.19052. [DOI] [PubMed] [Google Scholar]
  21. Pikaar J. C., Voorhout W. F., van Golde L. M., Verhoef J., Van Strijp J. A., van Iwaarden J. F. Opsonic activities of surfactant proteins A and D in phagocytosis of gram-negative bacteria by alveolar macrophages. J Infect Dis. 1995 Aug;172(2):481–489. doi: 10.1093/infdis/172.2.481. [DOI] [PubMed] [Google Scholar]
  22. Rust K., Grosso L., Zhang V., Chang D., Persson A., Longmore W., Cai G. Z., Crouch E. Human surfactant protein D: SP-D contains a C-type lectin carbohydrate recognition domain. Arch Biochem Biophys. 1991 Oct;290(1):116–126. doi: 10.1016/0003-9861(91)90597-c. [DOI] [PubMed] [Google Scholar]
  23. Schelenz S., Malhotra R., Sim R. B., Holmskov U., Bancroft G. J. Binding of host collectins to the pathogenic yeast Cryptococcus neoformans: human surfactant protein D acts as an agglutinin for acapsular yeast cells. Infect Immun. 1995 Sep;63(9):3360–3366. doi: 10.1128/iai.63.9.3360-3366.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Schweinle J. E., Ezekowitz R. A., Tenner A. J., Kuhlman M., Joiner K. A. Human mannose-binding protein activates the alternative complement pathway and enhances serum bactericidal activity on a mannose-rich isolate of Salmonella. J Clin Invest. 1989 Dec;84(6):1821–1829. doi: 10.1172/JCI114367. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Sheriff S., Chang C. Y., Ezekowitz R. A. Human mannose-binding protein carbohydrate recognition domain trimerizes through a triple alpha-helical coiled-coil. Nat Struct Biol. 1994 Nov;1(11):789–794. doi: 10.1038/nsb1194-789. [DOI] [PubMed] [Google Scholar]
  26. Spissinger T., Schäfer K. P., Voss T. Assembly of the surfactant protein SP-A. Deletions in the globular domain interfere with the correct folding of the molecule. Eur J Biochem. 1991 Jul 1;199(1):65–71. doi: 10.1111/j.1432-1033.1991.tb16092.x. [DOI] [PubMed] [Google Scholar]
  27. Wakamiya N., Okuno Y., Sasao F., Ueda S., Yoshimatsu K., Naiki M., Kurimura T. Isolation and characterization of conglutinin as an influenza A virus inhibitor. Biochem Biophys Res Commun. 1992 Sep 30;187(3):1270–1278. doi: 10.1016/0006-291x(92)90440-v. [DOI] [PubMed] [Google Scholar]
  28. Weaver T. E., Whitsett J. A. Function and regulation of expression of pulmonary surfactant-associated proteins. Biochem J. 1991 Jan 15;273(Pt 2):249–264. doi: 10.1042/bj2730249. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. van Golde L. M. Potential role of surfactant proteins A and D in innate lung defense against pathogens. Biol Neonate. 1995;67 (Suppl 1):2–17. doi: 10.1159/000244202. [DOI] [PubMed] [Google Scholar]

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