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PI-PLC releases a 25-40 kDa protein cluster from the hamster oolemma and affects the sperm penetration assay

Coonrod, S; Naaby-Hansen, S; Shetty, J; Herr, J; (1999) PI-PLC releases a 25-40 kDa protein cluster from the hamster oolemma and affects the sperm penetration assay. Molecular Human Reproduction , 5 (11) 1027 - 1033.

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Abstract

The effects of phosphatidylinositol-specific phospholipase C (PI-PLC) on human sperm-hamster oocyte interaction were investigated to determine whether PI-PLC cleavable glycosylphosphatidyinositol (GPI)- anchored proteins are involved in sperm-egg binding and fusion. Two- dimensional electrophoresis was then utilized to visualize proteins released from hamster oocytes following PI-PLC treatment. For the binding and fusion assay, either spermatozoa or eggs were treated with 1 IU/ml PI-PLC for 30 min and washed prior to gamete co-incubation. Treatment of human spermatozoa with PI-PLC significantly (P </= 0.05) enhanced sperm-egg binding while having no effect on sperm-egg fusion. Treatment of zona-free hamster oocytes with PI-PLC blocked sperm-egg binding and fusion. In order to identify the oolemmal GPI-anchored proteins involved in fertilization, egg surface proteins were labelled with sulpho-NHS biotin and either mock treated or treated with PI-PLC. Egg protein extracts and egg supernatant proteins from each group were then analysed by two-dimensional gel electrophoresis followed by avidin blotting. Comparison of blots demonstrated that a predominant biotinylated 25-40 kDa protein cluster (pI 5-6) apparent in the mock treated egg extract blot was absent in the PI-PLC treated egg extract blot. A protein cluster of identical molecular weight and isoelectric point as the predominant 25-40 kDa protein cluster was observed in the PI-PLC supernatant blot while no proteins could be seen in the control supernatant blot. These results demonstrate that treatment of hamster oocytes with PI-PLC inhibits sperm-egg interaction and releases a 25-40 kDa protein cluster (pI 5-6) from the oolemma. It is likely that this released protein cluster represents an oolemmal GPI-linked surface protein(s) which is involved in human sperm-hamster egg interaction

Type:Article
Title:PI-PLC releases a 25-40 kDa protein cluster from the hamster oolemma and affects the sperm penetration assay
Additional information:UI - 20009811 LA - Eng RN - EC 3.1.4.10 (1-phosphatidylinositol phosphodiesterase) RN - EC 3.1.4.3 (Phospholipase C) RN - 0 (Egg Proteins) PT - JOURNAL ARTICLE ID - U54 HD 29099/HD/NICHD ID -P30-28934 ID - P32-DK07642/DK/NIDDK ID - + DA - 19991130 IS -1360-9947 SB - M CY - ENGLAND JC - CWO AA - Author EM - 200002
Keywords:release, Proteins, hamster, sperm, egg, 1 phosphatidylinositol phosphodiesterase, Animal, Biotin, Biotinylation, drug effects, Egg Proteins, Electrophoresis, Gel, Two-Dimensional, Female, Hamsters, Isoelectric Point, Male, metabolism, Molecular Weight, oocytes, Ovum, pharmacology, Phospholipase C, physiology, Sequence Analysis, Sperm-Ovum Interactions, Spermatozoa, Support, Non-U.S.Gov't, Support, U.S.Gov't, P.H.S., eggs, fertilization, control, ANS
UCL classification:UCL > School of Life and Medical Sciences > Faculty of Life Sciences > Biosciences (Division of)

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