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. 1993 Apr 15;291(Pt 2):601–608. doi: 10.1042/bj2910601

Isolation, characterization and localization of annexin V from chicken liver.

C M Boustead 1, R Brown 1, J H Walker 1
PMCID: PMC1132566  PMID: 8484740

Abstract

Annexin V has been purified from chicken liver; 40 mg of annexin V was obtained per kg of tissue. In contrast with mammalian liver, very little annexin VI was obtained. Surprisingly, chicken liver annexin V resembles mammalian annexin IV in its M(r) (32,500) and its isoelectric point (5.6), but amino-acid-sequence analysis demonstrates identity with chicken annexin V (anchorin CII). It binds to phospholipids in a Ca(2+)-dependent manner with free-Ca2+ concentrations for half-maximal binding to phosphatidylserine and phosphatidic acid of 10 microM; phosphatidylethanolamine of 32 microM and phosphatidylinositol of 90 microM. No binding to phosphatidylcholine was observed at Ca2+ concentrations up to 300 microM. In isolated liver membranes a significant proportion of annexin V was not extractable with EGTA but could only be extracted with Triton X-100, suggesting the existence of a tightly membrane-associated form of annexin V. A specific antiserum to chicken annexin V was used to localize the protein in adult and embryonic chicken liver. In the adult, annexin V was highly concentrated in epithelial cells lining the bile ducts, and along the bile canaliculi. In embryonic liver, strong staining of the bile-duct epithelial cells was again evident, and in addition, endothelial cells were strongly immunoreactive.

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Selected References

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

  1. Anderson L., Anderson N. G. High resolution two-dimensional electrophoresis of human plasma proteins. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5421–5425. doi: 10.1073/pnas.74.12.5421. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Andree H. A., Reutelingsperger C. P., Hauptmann R., Hemker H. C., Hermens W. T., Willems G. M. Binding of vascular anticoagulant alpha (VAC alpha) to planar phospholipid bilayers. J Biol Chem. 1990 Mar 25;265(9):4923–4928. [PubMed] [Google Scholar]
  3. Bianchi R., Giambanco I., Ceccarelli P., Pula G., Donato R. Membrane-bound annexin V isoforms (CaBP33 and CaBP37) and annexin VI in bovine tissues behave like integral membrane proteins. FEBS Lett. 1992 Jan 20;296(2):158–162. doi: 10.1016/0014-5793(92)80369-r. [DOI] [PubMed] [Google Scholar]
  4. Boustead C. M., Walker J. H., Geisow M. J. Isolation and characterization of two novel calcium-dependent phospholipid-binding proteins from bovine lung. FEBS Lett. 1988 Jun 20;233(2):233–238. doi: 10.1016/0014-5793(88)80433-2. [DOI] [PubMed] [Google Scholar]
  5. Boustead C. M., Walker J. H., Kennedy D., Waller D. A. Crystallization and preliminary X-ray studies of annexin IV (endonexin), a calcium-dependent phospholipid-binding protein. FEBS Lett. 1991 Feb 25;279(2):187–189. doi: 10.1016/0014-5793(91)80145-s. [DOI] [PubMed] [Google Scholar]
  6. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  7. 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]
  8. Campos-Gonzalez R., Kanemitsu M., Boynton A. L. Epidermal growth factor induces the accumulation of calpactin II on the cell surface during membrane ruffling. Cell Motil Cytoskeleton. 1990;15(1):34–40. doi: 10.1002/cm.970150106. [DOI] [PubMed] [Google Scholar]
  9. Christmas P., Callaway J., Fallon J., Jones J., Haigler H. T. Selective secretion of annexin 1, a protein without a signal sequence, by the human prostate gland. J Biol Chem. 1991 Feb 5;266(4):2499–2507. [PubMed] [Google Scholar]
  10. 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]
  11. 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]
  12. Denton R. M., Richards D. A., Chin J. G. Calcium ions and the regulation of NAD+-linked isocitrate dehydrogenase from the mitochondria of rat heart and other tissues. Biochem J. 1978 Dec 15;176(3):899–906. doi: 10.1042/bj1760899. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Ernst J. D. Annexin III translocates to the periphagosomal region when neutrophils ingest opsonized yeast. J Immunol. 1991 May 1;146(9):3110–3114. [PubMed] [Google Scholar]
  14. Feldman H., Rodbard D., Levine D. Mathematical theory of cross-reactive radioimmunoassay and ligand-binding systems of equilibrium. Anal Biochem. 1972 Feb;45(2):530–556. doi: 10.1016/0003-2697(72)90216-3. [DOI] [PubMed] [Google Scholar]
  15. Fernandez M. P., Selmin O., Martin G. R., Yamada Y., Pfäffle M., Deutzmann R., Mollenhauer J., von der Mark K. The structure of anchorin CII, a collagen binding protein isolated from chondrocyte membrane. J Biol Chem. 1990 May 15;265(14):8344–8344. [PubMed] [Google Scholar]
  16. Gerke V. Identification of a homologue for annexin VII (synexin) in Dictyostelium discoideum. J Biol Chem. 1991 Jan 25;266(3):1697–1700. [PubMed] [Google Scholar]
  17. Giambanco I., Pula G., Ceccarelli P., Bianchi R., Donato R. Immunohistochemical localization of annexin V (CaBP33) in rat organs. J Histochem Cytochem. 1991 Sep;39(9):1189–1198. doi: 10.1177/39.9.1833446. [DOI] [PubMed] [Google Scholar]
  18. Greenberg M. E., Brackenbury R., Edelman G. M. Changes in the distribution of the 34-kdalton tyrosine kinase substrate during differentiation and maturation of chicken tissues. J Cell Biol. 1984 Feb;98(2):473–486. doi: 10.1083/jcb.98.2.473. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Greenwood M., Tsang A. Sequence and expression of annexin VII of Dictyostelium discoideum. Biochim Biophys Acta. 1991 Mar 26;1088(3):429–432. doi: 10.1016/0167-4781(91)90137-b. [DOI] [PubMed] [Google Scholar]
  20. Horseman N. D. A prolactin-inducible gene product which is a member of the calpactin/lipocortin family. Mol Endocrinol. 1989 May;3(5):773–779. doi: 10.1210/mend-3-5-773. [DOI] [PubMed] [Google Scholar]
  21. Huber R., Berendes R., Burger A., Schneider M., Karshikov A., Luecke H., Römisch J., Paques E. Crystal and molecular structure of human annexin V after refinement. Implications for structure, membrane binding and ion channel formation of the annexin family of proteins. J Mol Biol. 1992 Feb 5;223(3):683–704. doi: 10.1016/0022-2836(92)90984-r. [DOI] [PubMed] [Google Scholar]
  22. Huber R., Römisch J., Paques E. P. The crystal and molecular structure of human annexin V, an anticoagulant protein that binds to calcium and membranes. EMBO J. 1990 Dec;9(12):3867–3874. doi: 10.1002/j.1460-2075.1990.tb07605.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Huber R., Schneider M., Mayr I., Römisch J., Paques E. P. The calcium binding sites in human annexin V by crystal structure analysis at 2.0 A resolution. Implications for membrane binding and calcium channel activity. FEBS Lett. 1990 Nov 26;275(1-2):15–21. doi: 10.1016/0014-5793(90)81428-q. [DOI] [PubMed] [Google Scholar]
  24. 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]
  25. 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]
  26. Johnston P. A., Perin M. S., Reynolds G. A., Wasserman S. A., Südhof T. C. Two novel annexins from Drosophila melanogaster. Cloning, characterization, and differential expression in development. J Biol Chem. 1990 Jul 5;265(19):11382–11388. [PubMed] [Google Scholar]
  27. Khanna N. C., Helwig E. D., Ikebuchi N. W., Fitzpatrick S., Bajwa R., Waisman D. M. Purification and characterization of annexin proteins from bovine lung. Biochemistry. 1990 May 22;29(20):4852–4862. doi: 10.1021/bi00472a015. [DOI] [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. Kobayashi R., Tashima Y. Purification, biological properties and partial sequence analysis of 67-kDa calcimedin and its 34-kDa fragment from chicken gizzard. Eur J Biochem. 1990 Mar 10;188(2):447–453. doi: 10.1111/j.1432-1033.1990.tb15422.x. [DOI] [PubMed] [Google Scholar]
  30. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  31. Lowe P. J., Miyai K., Steinbach J. H., Hardison W. G. Hormonal regulation of hepatocyte tight junctional permeability. Am J Physiol. 1988 Oct;255(4 Pt 1):G454–G461. doi: 10.1152/ajpgi.1988.255.4.G454. [DOI] [PubMed] [Google Scholar]
  32. 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]
  33. Nathanson M. H., Gautam A., Bruck R., Isales C. M., Boyer J. L. Effects of Ca2+ agonists on cytosolic Ca2+ in isolated hepatocytes and on bile secretion in the isolated perfused rat liver. Hepatology. 1992 Jan;15(1):107–116. doi: 10.1002/hep.1840150119. [DOI] [PubMed] [Google Scholar]
  34. O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
  35. Owens R. J., Crumpton M. J. Isolation and characterization of a novel 68,000-Mr Ca2+-binding protein of lymphocyte plasma membrane. Biochem J. 1984 Apr 1;219(1):309–316. doi: 10.1042/bj2190309. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Pepinsky R. B., Sinclair L. K. Epidermal growth factor-dependent phosphorylation of lipocortin. Nature. 1986 May 1;321(6065):81–84. doi: 10.1038/321081a0. [DOI] [PubMed] [Google Scholar]
  37. Pfäffle M., Ruggiero F., Hofmann H., Fernández M. P., Selmin O., Yamada Y., Garrone R., von der Mark K. Biosynthesis, secretion and extracellular localization of anchorin CII, a collagen-binding protein of the calpactin family. EMBO J. 1988 Aug;7(8):2335–2342. doi: 10.1002/j.1460-2075.1988.tb03077.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Pilar Fernandez M., Selmin O., Martin G. R., Yamada Y., Pfäffle M., Deutzmann R., Mollenhauer J., von der Mark K. The structure of anchorin CII, a collagen binding protein isolated from chondrocyte membrane. J Biol Chem. 1988 Apr 25;263(12):5921–5925. [PubMed] [Google Scholar]
  39. Reeves J. P., Dowben R. M. Formation and properties of thin-walled phospholipid vesicles. J Cell Physiol. 1969 Feb;73(1):49–60. doi: 10.1002/jcp.1040730108. [DOI] [PubMed] [Google Scholar]
  40. Robitzki A., Schröder H. C., Ugarković D., Gramzow M., Fritsche U., Batel R., Müller W. E. cDNA structure and expression of calpactin, a peptide involved in Ca2(+)-dependent cell aggregation in sponges. Biochem J. 1990 Oct 15;271(2):415–420. doi: 10.1042/bj2710415. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Rojas E., Pollard H. B., Haigler H. T., Parra C., Burns A. L. Calcium-activated endonexin II forms calcium channels across acidic phospholipid bilayer membranes. J Biol Chem. 1990 Dec 5;265(34):21207–21215. [PubMed] [Google Scholar]
  42. Römisch J., Pâques E. P. Annexins: calcium-binding proteins of multi-functional importance? Med Microbiol Immunol. 1991;180(3):109–126. doi: 10.1007/BF00206115. [DOI] [PubMed] [Google Scholar]
  43. Römisch J., Schorlemmer U., Fickenscher K., Pâques E. P., Heimburger N. Anticoagulant properties of placenta protein 4 (annexin V). Thromb Res. 1990 Dec 1;60(5):355–366. doi: 10.1016/0049-3848(90)90218-2. [DOI] [PubMed] [Google Scholar]
  44. 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]
  45. Sato E. F., Morimoto Y. M., Matsuno T., Miyahara M., Utsumi K. Neutrophil specific 33 kDa protein: its Ca2+- and phospholipid-dependent intracellular translocation. FEBS Lett. 1987 Apr 6;214(1):181–186. doi: 10.1016/0014-5793(87)80038-8. [DOI] [PubMed] [Google Scholar]
  46. Schlaepfer D. D., Fisher D. A., Brandt M. E., Bode H. R., Jones J. M., Haigler H. T. Identification of a novel annexin in Hydra vulgaris. Characterization, cDNA cloning, and protein kinase C phosphorylation of annexin XII. J Biol Chem. 1992 May 15;267(14):9529–9539. [PubMed] [Google Scholar]
  47. Schlaepfer D. D., Jones J., Haigler H. T. Inhibition of protein kinase C by annexin V. Biochemistry. 1992 Feb 18;31(6):1886–1891. doi: 10.1021/bi00121a043. [DOI] [PubMed] [Google Scholar]
  48. Smith P. K., Krohn R. I., Hermanson G. T., Mallia A. K., Gartner F. H., Provenzano M. D., Fujimoto E. K., Goeke N. M., Olson B. J., Klenk D. C. Measurement of protein using bicinchoninic acid. Anal Biochem. 1985 Oct;150(1):76–85. doi: 10.1016/0003-2697(85)90442-7. [DOI] [PubMed] [Google Scholar]
  49. Südhof T. C., Ebbecke M., Walker J. H., Fritsche U., Boustead C. Isolation of mammalian calelectrins: a new class of ubiquitous Ca2+-regulated proteins. Biochemistry. 1984 Mar 13;23(6):1103–1109. doi: 10.1021/bi00301a010. [DOI] [PubMed] [Google Scholar]
  50. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Walker J. H. Isolation from cholinergic synapses of a protein that binds to membranes in a calcium-dependent manner. J Neurochem. 1982 Sep;39(3):815–823. doi: 10.1111/j.1471-4159.1982.tb07965.x. [DOI] [PubMed] [Google Scholar]
  52. Walker J. H., Kristjansson G. I., Stadler H. Identification of a synaptic vesicle antigen (Mr 86,000) conserved between Torpedo and rat. J Neurochem. 1986 Mar;46(3):875–881. doi: 10.1111/j.1471-4159.1986.tb13053.x. [DOI] [PubMed] [Google Scholar]
  53. Watanabe S., Smith C. R., Phillips M. J. Coordination of the contractile activity of bile canaliculi. Evidence from calcium microinjection of triplet hepatocytes. Lab Invest. 1985 Sep;53(3):275–279. [PubMed] [Google Scholar]
  54. Woolgar J. A., Boustead C. M., Walker J. H. Characterization of annexins in mammalian brain. J Neurochem. 1990 Jan;54(1):62–71. doi: 10.1111/j.1471-4159.1990.tb13283.x. [DOI] [PubMed] [Google Scholar]
  55. von der Mark K., Hofmann C., Pfäffle M., Gropp R. Anchorin CII sequence now corrected. Trends Biochem Sci. 1990 Jan;15(1):12–12. doi: 10.1016/0968-0004(90)90119-v. [DOI] [PubMed] [Google Scholar]

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