Isolation and characterization of a rabbit epididymal secretory glycoprotein that associates to the spermatozoon surface.
Using a combination of gel filtration and hydroxyapatite chromatography, a major secretory glycoprotein (EP140) was purified from rabbit epididymal fluid. The protein had an apparent Mr of approximately 140 kDa under native conditions but dissociated into 2 equimolar amounts of glycosylated subunits, alpha and beta of Mr 35 and 33 kDa, upon sodium dodecylsulfate polyacrylamide gel electrophoresis in absence of reducing agents. Thus EP140 appears to be a tetramer composed of 2 alpha and 2 beta subunits, held together by noncovalent forces. Proteolytic peptide mapping, amino acid analysis, and determination of partial amino acid sequences suggested that the 2 subunits were very similar, differing only at some punctual residues in their primary structures. The amino acid sequences obtained did not show significant similarity to any known protein. Western blot determinations with a specific antibody indicated that no EP140-immunorelated protein was detected either in testis or blood from the rabbit nor in epididymides from rats or hamsters. EP140 was shown to associated to the spermatozoon surface, mainly at the acrosomal zone and in the middle piece, and this association progressively increased during the transit of the spermatozoon through the epididymis.
Nieto
A
A
Centro de Biología Molecular Severo Ochoa, Facultad de Ciencias, Universidad Autónoma de Madrid, Cantoblanco, Spain.
Gutierrez Sagal
R
R
Pérez Martinez
M
M
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002850:Chromatography, Gel; D006224:Cricetinae; D004591:Electrophoresis, Polyacrylamide Gel; D004822:Epididymis / Q000378:metabolism*; D006023:Glycoproteins / Q000737:chemistry / Q000302:isolation & purification*; D036341:Intercellular Signaling Peptides and Proteins; D008297:Male; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010449:Peptide Mapping; D011817:Rabbits; D051381:Rats; D013094:Spermatozoa / Q000737:chemistry*; D013499:Surface Properties
D006023:Glycoproteins; D036341:Intercellular Signaling Peptides and Proteins; C107476:epididymal secretory glycoprotein, 140-kDa