title: Isolation of a calcium-binding protein of the acrosomal membrane of bovine spermatozoa.
abstract: The mammalian sperm acrosome reaction is a calcium-dependent exocytotic event characterized by extensive fusion between the plasma and the outer acrosomal membrane. The mechanisms by which elevation of cytosolic calcium initiates the membrane fusion process are not understood and the present study was undertaken to identify calcium-binding proteins in the acrosomal membrane (AM) of bovine spermatozoa. Sperm heads, purified from sonicated spermatozoa, were used to isolate an acrosomal membrane-enriched fraction on Percoll density gradients. Using SDS-PAGE and a (45)Ca(2+)-blot overlay assay, calcium-binding proteins of 64, 45, 43, and 39kDa were identified in the AM enriched fraction. Phase separation analysis with Triton X-114 identified the 64kDa polypeptide as an integral membrane protein. The 64kDa polypeptide was purified and utilized to prepare a polyclonal antiserum. Both light and electron microscopic immunocytochemistry demonstrated that the protein was distributed throughout all domains of the acrosomal membrane. These results identify a 64kDa calcium-binding integral membrane protein of the mammalian acrosome. Its potential function in calcium-dependent membrane fusion events of the acrosome reaction and in fertilization is discussed.
authors: Department of Chemistry and Physics, Fayetteville State University, Fayetteville, NC 28301, USA. snagdas@uncfsu.edu
authors: Subir K
authors: SK
authors: Nagdas
authors: Teresa
authors: T
authors: Buchanan
authors: Shaina
authors: S
authors: McCaskill
authors: Jared
authors: J
authors: Mackey
authors: George E
authors: GE
authors: Alvarez
authors: Samir
authors: S
authors: Raychoudhury
mesh_terms: D000177:Acrosome / Q000378:metabolism*; D000818:Animals; D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D002417:Cattle; D007425:Intracellular Membranes / Q000378:metabolism*; D008297:Male; D008970:Molecular Weight; D021381:Protein Transport
chemical_list: D002135:Calcium-Binding Proteins
references: Nature. 1970 Aug 15;227(5259):680-5
references: 5432063
references: Anal Biochem. 1987 Nov 1;166(2):308-12
references: 2449094
references: Nature. 1995 Jun 22;375(6533):645-53
references: 7791897
references: Dev Biol. 1988 Sep;129(1):12-24
references: 3137110
references: J Exp Zool. 1979 Apr;208(1):41-56
references: 381567
references: Biochim Biophys Acta. 1975 Nov 3;406(4):479-88
references: 1237320
references: Mol Reprod Dev. 1994 Dec;39(4):423-32
references: 7893491
references: Curr Opin Cell Biol. 1995 Oct;7(5):658-64
references: 8573340
references: J Exp Zool. 1986 Feb;237(2):257-62
references: 3950567
references: Biol Reprod. 1987 Dec;37(5):1249-58
references: 2832008
references: J Cell Biol. 1967 Sep;34(3):C1-5
references: 6050956
references: J Exp Zool. 1986 Apr;238(1):113-8
references: 3086485
references: Dev Biol. 1997 Feb 1;182(1):67-75
references: 9073447
references: J Biol Chem. 1989 Aug 15;264(23):13586-90
references: 2503502
references: Dev Biol. 1992 Aug;152(2):304-14
references: 1379559
references: Mol Reprod Dev. 1995 Aug;41(4):513-20
references: 7576619
references: Exp Cell Res. 1974 Nov;89(1):161-74
references: 4435057
references: J Biol Chem. 1994 Dec 23;269(51):32607-14
references: 7798265
references: Anal Biochem. 1976 May 7;72:248-54
references: 942051
references: Biol Reprod. 1996 Jan;54(1):111-21
references: 8838007
references: Exp Cell Res. 1975 Nov;96(1):23-30
references: 1193170
references: J Biol Chem. 1981 Feb 25;256(4):1604-7
references: 6257680
references: J Cell Biol. 1995 Aug;130(4):857-69
references: 7642703
references: J Cell Biol. 1996 Aug;134(3):637-45
references: 8707844
references: Gamete Res. 1988 Aug;20(4):397-411
references: 3235048
references: Mol Reprod Dev. 1994 Dec;39(4):433-8
references: 7893492
references: Mol Reprod Dev. 1995 Nov;42(3):325-33
references: 8579847
references: Trends Neurosci. 1995 Apr;18(4):191-6
references: 7778190
references: Dev Biol. 1994 Sep;165(1):152-64
references: 8088433
references: EMBO J. 2012 Apr 4;31(7):1654-65
references: 22354039
references: Science. 1996 Jul 12;273(5272):248-51
references: 8662510
references: Biochemistry. 1971 Jun 22;10(13):2606-17
references: 4326772
references: Reproduction. 2001 Dec;122(6):939-46
references: 11732989
references: J Immunol Methods. 1984 Sep 4;72(2):313-40
references: 6206159
references: J Biol Chem. 2009 Apr 17;284(16):10491-503
references: 19208619
references: J Exp Zool. 1993 Nov 1;267(3):350-5
references: 8228870
references: Anal Biochem. 1981 Nov 15;118(1):197-203
references: 6175245
references: Mol Reprod Dev. 1996 Oct;45(2):204-11
references: 8914078
references: Proc Natl Acad Sci U S A. 1996 Sep 17;93(19):10471-6
references: 8816825
references: Science. 1992 May 15;256(5059):1021-5
references: 1589771
references: Methods Enzymol. 1986;134:467-72
references: 3547038
references: Dev Biol. 1989 Sep;135(1):133-46
references: 2504631
references: Biol Reprod. 1985 Aug;33(1):235-46
references: 3840700
references: Biol Reprod. 2002 Jan;66(1):50-6
references: 11751263
references: Science. 1987 Jan 30;235(4788):553-60
references: 3027891
references: J Biochem. 1984 Feb;95(2):511-9
references: 6715311
references: J Ultrastruct Res. 1985 Jan;90(1):9-25
references: 2415711
references: Mol Reprod Dev. 1993 Nov;36(3):368-76
references: 8286120
references: Science. 1995 Apr 14;268(5208):221-5
references: 7716512
references: Biol Reprod. 1997 Mar;56(3):569-74
references: 9046998
references: Biol Reprod. 1995 Feb;52(2):373-81
references: 7536051
references: Commun Integr Biol. 2012 Jan 1;5(1):68-70
references: 22482014
references: J Biol Chem. 2010 Aug 20;285(34):26269-78
references: 20551332
references: J Cell Biol. 1983 Oct;97(4):1277-82
references: 6413513
references: Mol Reprod Dev. 1994 Nov;39(3):297-308
references: 7888169
references: Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350-4
references: 388439
references: J Biol Chem. 1994 Sep 23;269(38):23583-9
references: 8089126
references: Endocrinology. 1996 Sep;137(9):3999-4009
references: 8756577
references: J Biol Chem. 1994 May 6;269(18):13254-8
references: 8175755