Skip to main content
PMC home page
The EMBO Journal logoLink to The EMBO Journal
. 1990 Dec;9(12):3867–3874. doi: 10.1002/j.1460-2075.1990.tb07605.x

The crystal and molecular structure of human annexin V, an anticoagulant protein that binds to calcium and membranes.

R Huber 1, J Römisch 1, E P Paques 1
PMCID: PMC552154  PMID: 2147412

Abstract

Human annexin V (PP4), a member of the family of calcium, membrane binding proteins, has been crystallized in the presence of calcium and analysed by crystallography by multiple isomorphic replacement at 3 A and preliminarily refined at 2.5 A resolution. The molecule has dimensions of 64 x 40 x 30 A3 and is folded into four domains of similar structure. Each domain consists of five alpha-helices wound into a right-handed superhelix yielding a globular structure of approximately 18 A diameter. The domains have hydrophobic cores whose amino acid sequences are conserved between the domains and within the annexin family of proteins. The four domains are folded into an almost planar array by tight (hydrophobic) pair-wise packing of domains II and III and I and IV to generate modules (II-III) and (I-IV), respectively. The assembly is symmetric with three parallel approximate diads relating II to III, I to IV and the module (II-III) to (I-IV), respectively. The latter diad marks a channel through the centre of the molecule coated with charged amino acid residues. The protein has structural features of channel forming membrane proteins and a polar surface characteristic of soluble proteins. It is a member of the third class of amphipathic proteins different from soluble and membrane proteins.

Full text

PDF
3868

Images in this article

3868Fig. 1.
on p.3868

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ando Y., Imamura S., Hong Y. M., Owada M. K., Kakunaga T., Kannagi R. Enhancement of calcium sensitivity of lipocortin I in phospholipid binding induced by limited proteolysis and phosphorylation at the amino terminus as analyzed by phospholipid affinity column chromatography. J Biol Chem. 1989 Apr 25;264(12):6948–6955. [PubMed] [Google Scholar]
  2. Bode W., Mayr I., Baumann U., Huber R., Stone S. R., Hofsteenge J. The refined 1.9 A crystal structure of human alpha-thrombin: interaction with D-Phe-Pro-Arg chloromethylketone and significance of the Tyr-Pro-Pro-Trp insertion segment. EMBO J. 1989 Nov;8(11):3467–3475. doi: 10.1002/j.1460-2075.1989.tb08511.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Brandhuber B. J., Boone T., Kenney W. C., McKay D. B. Three-dimensional structure of interleukin-2. Science. 1987 Dec 18;238(4834):1707–1709. doi: 10.1126/science.3500515. [DOI] [PubMed] [Google Scholar]
  4. Burgoyne R. D., Geisow M. J. The annexin family of calcium-binding proteins. Review article. Cell Calcium. 1989 Jan;10(1):1–10. doi: 10.1016/0143-4160(89)90038-9. [DOI] [PubMed] [Google Scholar]
  5. Burns A. L., Magendzo K., Shirvan A., Srivastava M., Rojas E., Alijani M. R., Pollard H. B. Calcium channel activity of purified human synexin and structure of the human synexin gene. Proc Natl Acad Sci U S A. 1989 May;86(10):3798–3802. doi: 10.1073/pnas.86.10.3798. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Creutz C. E., Zaks W. J., Hamman H. C., Crane S., Martin W. H., Gould K. L., Oddie K. M., Parsons S. J. Identification of chromaffin granule-binding proteins. Relationship of the chromobindins to calelectrin, synhibin, and the tyrosine kinase substrates p35 and p36. J Biol Chem. 1987 Feb 5;262(4):1860–1868. [PubMed] [Google Scholar]
  7. Crompton M. R., Moss S. E., Crumpton M. J. Diversity in the lipocortin/calpactin family. Cell. 1988 Oct 7;55(1):1–3. doi: 10.1016/0092-8674(88)90002-5. [DOI] [PubMed] [Google Scholar]
  8. Crompton M. R., Owens R. J., Totty N. F., Moss S. E., Waterfield M. D., Crumpton M. J. Primary structure of the human, membrane-associated Ca2+-binding protein p68 a novel member of a protein family. EMBO J. 1988 Jan;7(1):21–27. doi: 10.1002/j.1460-2075.1988.tb02779.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Crumpton M. J., Dedman J. R. Protein terminology tangle. Nature. 1990 May 17;345(6272):212–212. doi: 10.1038/345212a0. [DOI] [PubMed] [Google Scholar]
  10. Declercq J. P., Tinant B., Parello J., Etienne G., Huber R. Crystal structure determination and refinement of pike 4.10 parvalbumin (minor component from Esox lucius). J Mol Biol. 1988 Jul 20;202(2):349–353. doi: 10.1016/0022-2836(88)90464-0. [DOI] [PubMed] [Google Scholar]
  11. Drust D. S., Creutz C. E. Aggregation of chromaffin granules by calpactin at micromolar levels of calcium. Nature. 1988 Jan 7;331(6151):88–91. doi: 10.1038/331088a0. [DOI] [PubMed] [Google Scholar]
  12. Flaherty M. J., West S., Heimark R. L., Fujikawa K., Tait J. F. Placental anticoagulant protein-I: measurement in extracellular fluids and cells of the hemostatic system. J Lab Clin Med. 1990 Feb;115(2):174–181. [PubMed] [Google Scholar]
  13. Geisow M. J., Fritsche U., Hexham J. M., Dash B., Johnson T. A consensus amino-acid sequence repeat in Torpedo and mammalian Ca2+-dependent membrane-binding proteins. Nature. 1986 Apr 17;320(6063):636–638. doi: 10.1038/320636a0. [DOI] [PubMed] [Google Scholar]
  14. Goulding N. J., Godolphin J. L., Sharland P. R., Peers S. H., Sampson M., Maddison P. J., Flower R. J. Anti-inflammatory lipocortin 1 production by peripheral blood leucocytes in response to hydrocortisone. Lancet. 1990 Jun 16;335(8703):1416–1418. doi: 10.1016/0140-6736(90)91445-g. [DOI] [PubMed] [Google Scholar]
  15. Grundmann U., Abel K. J., Bohn H., Löbermann H., Lottspeich F., Küpper H. Characterization of cDNA encoding human placental anticoagulant protein (PP4): homology with the lipocortin family. Proc Natl Acad Sci U S A. 1988 Jun;85(11):3708–3712. doi: 10.1073/pnas.85.11.3708. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Herzberg O., James M. N. Structure of the calcium regulatory muscle protein troponin-C at 2.8 A resolution. Nature. 1985 Feb 21;313(6004):653–659. doi: 10.1038/313653a0. [DOI] [PubMed] [Google Scholar]
  17. Hirata F., Schiffmann E., Venkatasubramanian K., Salomon D., Axelrod J. A phospholipase A2 inhibitory protein in rabbit neutrophils induced by glucocorticoids. Proc Natl Acad Sci U S A. 1980 May;77(5):2533–2536. doi: 10.1073/pnas.77.5.2533. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hullin F., Raynal P., Ragab-Thomas J. M., Fauvel J., Chap H. Effect of dexamethasone on prostaglandin synthesis and on lipocortin status in human endothelial cells. Inhibition of prostaglandin I2 synthesis occurring without alteration of arachidonic acid liberation and of lipocortin synthesis. J Biol Chem. 1989 Feb 25;264(6):3506–3513. [PubMed] [Google Scholar]
  19. Isacke C. M., Lindberg R. A., Hunter T. Synthesis of p36 and p35 is increased when U-937 cells differentiate in culture but expression is not inducible by glucocorticoids. Mol Cell Biol. 1989 Jan;9(1):232–240. doi: 10.1128/mcb.9.1.232. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Iwasaki A., Suda M., Nakao H., Nagoya T., Saino Y., Arai K., Mizoguchi T., Sato F., Yoshizaki H., Hirata M. Structure and expression of cDNA for an inhibitor of blood coagulation isolated from human placenta: a new lipocortin-like protein. J Biochem. 1987 Nov;102(5):1261–1273. doi: 10.1093/oxfordjournals.jbchem.a122165. [DOI] [PubMed] [Google Scholar]
  21. Johnsson N., Johnsson K., Weber K. A discontinuous epitope on p36, the major substrate of src tyrosine-protein-kinase, brings the phosphorylation site into the neighbourhood of a consensus sequence for Ca2+/lipid-binding proteins. FEBS Lett. 1988 Aug 15;236(1):201–204. doi: 10.1016/0014-5793(88)80314-4. [DOI] [PubMed] [Google Scholar]
  22. Johnsson N., Marriott G., Weber K. p36, the major cytoplasmic substrate of src tyrosine protein kinase, binds to its p11 regulatory subunit via a short amino-terminal amphiphatic helix. EMBO J. 1988 Aug;7(8):2435–2442. doi: 10.1002/j.1460-2075.1988.tb03089.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Johnsson N., Vandekerckhove J., Van Damme J., Weber K. Binding sites for calcium, lipid and p11 on p36, the substrate of retroviral tyrosine-specific protein kinases. FEBS Lett. 1986 Mar 31;198(2):361–364. doi: 10.1016/0014-5793(86)80437-9. [DOI] [PubMed] [Google Scholar]
  24. Johnsson N., Weber K. Alkylation of cysteine 82 of p11 abolishes the complex formation with the tyrosine-protein kinase substrate p36 (annexin 2, calpactin 1, lipocortin 2). J Biol Chem. 1990 Aug 25;265(24):14464–14468. [PubMed] [Google Scholar]
  25. Johnsson N., Weber K. Structural analysis of p36, a Ca2+/lipid-binding protein of the annexin family, by proteolysis and chemical fragmentation. Eur J Biochem. 1990 Feb 22;188(1):1–7. doi: 10.1111/j.1432-1033.1990.tb15363.x. [DOI] [PubMed] [Google Scholar]
  26. Kaetzel M. A., Dedman J. R. Affinity-purified site-directed antibody recognizes the entire annexin protein family. Biochem Biophys Res Commun. 1989 May 15;160(3):1233–1237. doi: 10.1016/s0006-291x(89)80135-4. [DOI] [PubMed] [Google Scholar]
  27. Kaplan R., Jaye M., Burgess W. H., Schlaepfer D. D., Haigler H. T. Cloning and expression of cDNA for human endonexin II, a Ca2+ and phospholipid binding protein. J Biol Chem. 1988 Jun 15;263(17):8037–8043. [PubMed] [Google Scholar]
  28. Klee C. B. Ca2+-dependent phospholipid- (and membrane-) binding proteins. Biochemistry. 1988 Sep 6;27(18):6645–6653. doi: 10.1021/bi00418a001. [DOI] [PubMed] [Google Scholar]
  29. Lewit-Bentley A., Doublié S., Fourme R., Bodo G. Crystallization and preliminary X-ray studies of human vascular anticoagulant protein. J Mol Biol. 1989 Dec 20;210(4):875–876. doi: 10.1016/0022-2836(89)90115-0. [DOI] [PubMed] [Google Scholar]
  30. Marriott G., Kirk W. R., Johnsson N., Weber K. Absorption and fluorescence spectroscopic studies of the Ca2(+)-dependent lipid binding protein p36: the annexin repeat as the Ca2+ binding site. Biochemistry. 1990 Jul 31;29(30):7004–7011. doi: 10.1021/bi00482a008. [DOI] [PubMed] [Google Scholar]
  31. Maurer-Fogy I., Reutelingsperger C. P., Pieters J., Bodo G., Stratowa C., Hauptmann R. Cloning and expression of cDNA for human vascular anticoagulant, a Ca2+-dependent phospholipid-binding protein. Eur J Biochem. 1988 Jul 1;174(4):585–592. doi: 10.1111/j.1432-1033.1988.tb14139.x. [DOI] [PubMed] [Google Scholar]
  32. Mitchell M. D., Lytton F. D., Varticovski L. Paradoxical stimulation of both lipocortin and prostaglandin production in human amnion cells by dexamethasone. Biochem Biophys Res Commun. 1988 Feb 29;151(1):137–141. doi: 10.1016/0006-291x(88)90569-4. [DOI] [PubMed] [Google Scholar]
  33. Moss S. E., Crompton M. R., Crumpton M. J. Molecular cloning of murine p68, a Ca2+-binding protein of the lipocortin family. Eur J Biochem. 1988 Oct 15;177(1):21–27. doi: 10.1111/j.1432-1033.1988.tb14340.x. [DOI] [PubMed] [Google Scholar]
  34. Newman R., Tucker A., Ferguson C., Tsernoglou D., Leonard K., Crumpton M. J. Crystallization of p68 on lipid monolayers and as three-dimensional single crystals. J Mol Biol. 1989 Mar 5;206(1):213–219. doi: 10.1016/0022-2836(89)90534-2. [DOI] [PubMed] [Google Scholar]
  35. Parker M. W., Pattus F., Tucker A. D., Tsernoglou D. Structure of the membrane-pore-forming fragment of colicin A. Nature. 1989 Jan 5;337(6202):93–96. doi: 10.1038/337093a0. [DOI] [PubMed] [Google Scholar]
  36. Powell M. A., Glenney J. R. Regulation of calpactin I phospholipid binding by calpactin I light-chain binding and phosphorylation by p60v-src. Biochem J. 1987 Oct 15;247(2):321–328. doi: 10.1042/bj2470321. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Remington S., Wiegand G., Huber R. Crystallographic refinement and atomic models of two different forms of citrate synthase at 2.7 and 1.7 A resolution. J Mol Biol. 1982 Jun 15;158(1):111–152. doi: 10.1016/0022-2836(82)90452-1. [DOI] [PubMed] [Google Scholar]
  38. Reutelingsperger C. P., Hornstra G., Hemker H. C. Isolation and partial purification of a novel anticoagulant from arteries of human umbilical cord. Eur J Biochem. 1985 Sep 16;151(3):625–629. doi: 10.1111/j.1432-1033.1985.tb09150.x. [DOI] [PubMed] [Google Scholar]
  39. Ross T. S., Tait J. F., Majerus P. W. Identity of inositol 1,2-cyclic phosphate 2-phosphohydrolase with lipocortin III. Science. 1990 May 4;248(4955):605–607. doi: 10.1126/science.2159184. [DOI] [PubMed] [Google Scholar]
  40. Rydel T. J., Ravichandran K. G., Tulinsky A., Bode W., Huber R., Roitsch C., Fenton J. W., 2nd The structure of a complex of recombinant hirudin and human alpha-thrombin. Science. 1990 Jul 20;249(4966):277–280. doi: 10.1126/science.2374926. [DOI] [PubMed] [Google Scholar]
  41. Römisch J., Heimburger N. Purification and characterization of six annexins from human placenta. Biol Chem Hoppe Seyler. 1990 May;371(5):383–388. doi: 10.1515/bchm3.1990.371.1.383. [DOI] [PubMed] [Google Scholar]
  42. Saris C. J., Tack B. F., Kristensen T., Glenney J. R., Jr, Hunter T. The cDNA sequence for the protein-tyrosine kinase substrate p36 (calpactin I heavy chain) reveals a multidomain protein with internal repeats. Cell. 1986 Jul 18;46(2):201–212. doi: 10.1016/0092-8674(86)90737-3. [DOI] [PubMed] [Google Scholar]
  43. Seaton B. A., Head J. F., Kaetzel M. A., Dedman J. R. Purification, crystallization, and preliminary X-ray diffraction analysis of rat kidney annexin V, a calcium-dependent phospholipid-binding protein. J Biol Chem. 1990 Mar 15;265(8):4567–4569. [PubMed] [Google Scholar]
  44. Smith V. L., Kaetzel M. A., Dedman J. R. Stimulus-response coupling: the search for intracellular calcium mediator proteins. Cell Regul. 1990 Jan;1(2):165–172. doi: 10.1091/mbc.1.2.165. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Szebenyi D. M., Moffat K. The refined structure of vitamin D-dependent calcium-binding protein from bovine intestine. Molecular details, ion binding, and implications for the structure of other calcium-binding proteins. J Biol Chem. 1986 Jul 5;261(19):8761–8777. [PubMed] [Google Scholar]
  46. Tait J. F., Gibson D., Fujikawa K. Phospholipid binding properties of human placental anticoagulant protein-I, a member of the lipocortin family. J Biol Chem. 1989 May 15;264(14):7944–7949. [PubMed] [Google Scholar]
  47. Tait J. F., Sakata M., McMullen B. A., Miao C. H., Funakoshi T., Hendrickson L. E., Fujikawa K. Placental anticoagulant proteins: isolation and comparative characterization four members of the lipocortin family. Biochemistry. 1988 Aug 23;27(17):6268–6276. doi: 10.1021/bi00417a011. [DOI] [PubMed] [Google Scholar]
  48. Taylor W. R., Geisow M. J. Predicted structure for the calcium-dependent membrane-binding proteins p35, p36, and p32. Protein Eng. 1987 Jun;1(3):183–187. doi: 10.1093/protein/1.3.183. [DOI] [PubMed] [Google Scholar]
  49. Tufty R. M., Kretsinger R. H. Troponin and parvalbumin calcium binding regions predicted in myosin light chain and T4 lysozyme. Science. 1975 Jan 17;187(4172):167–169. doi: 10.1126/science.1111094. [DOI] [PubMed] [Google Scholar]
  50. Weber P. C., Salemme F. R. Structural and functional diversity in 4-alpha-helical proteins. Nature. 1980 Sep 4;287(5777):82–84. doi: 10.1038/287082a0. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES