Identification of goat sperm ecto-cyclic AMP independent protein kinase substrate localized on sperm outer surface
✍ Scribed by Arunima Maiti; K.P. Mishra; G.C. Majumder
- Publisher
- John Wiley and Sons
- Year
- 2004
- Tongue
- English
- Weight
- 296 KB
- Volume
- 92
- Category
- Article
- ISSN
- 0730-2312
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✦ Synopsis
Abstract
We have demonstrated the location of a cyclic AMP independent serine/threonine protein kinase (ecto‐CIK) on the outer surface of mature goat spermatozoa. We purified and characterized the major physiological protein substrate (MPS) of ecto‐CIK. ^32^P‐labeled membrane proteins phosphorylated by endogenous ecto‐CIK of intact cauda‐epididymal spermatozoa was solubilized with 1% Triton X‐100 and then fractionated by following several chromatographic techniques like Sephacryl S‐300 molecular sieve chromatography, DEAE–cellulose ion‐exchange chromatography and chromatofocussing. The MPS of ecto‐CIK has been purified to apparent homogeneity and it was found to be a monomeric protein of 100 kDa. Three isoforms of MPS have been found with pI of 6.37, 6.05, and 5.14 and all these isoforms served as the specific substrate of ecto‐CIK. The ecto‐kinase has nearly 30 times greater affinity for MPS as compared to casein the most potent exogenous protein substrate. Addition of MPS (pI 5.14) antibody caused head‐to‐head sperm agglutination. The Fv/Fab fragment of anti‐MPS caused significant inhibition of sperm motility. The data show that MPS is an ecto‐protein localized on the sperm head. MPS may thus play an important role for the regulation of sperm–egg interaction. © 2004 Wiley‐Liss, Inc.