
TY  - JOUR
A1  - Barua, Madhabi 
A1  - Ghosh, Anil K. 
A1  - Majumder, Gopal C. 
T1  - Partial purification and characterization of a phosphoprotein phosphatase from sperm plasma membrane
JF  - Reproduction, Fertility and Development
JO  - Reprod. Fertil. Dev.
Y1  - 2000
VL  - 11
IS  - 6
SP  - 379
EP  - 386
KW  - goat, external cell surface protein dephosphorylation, spermatozoa.
UR  - https://doi.org/10.1071/RD99095 
N2  - A phosphoprotein phosphatase (PPase M-I) that dephosphorylates serine and threonine residues of histones was isolated from the goat cauda-epididymal sperm plasma membrane and partially characterized. The PPase was solubilized from the sperm membrane by treating it with 0.1 N NaOH at pH 11.4 and the solubilized enzyme was partially purified by concanavalin A-sepharose affinity chromatography and high-performance liquid chromatography (HPLC), revealing it to be a 520-kDa protein. The PPase gave a single protein band in native polyacrylamide gel electrophoresis (PAGE), but in the presence of SDS it resolved into multiple proteins (35&#8211;170 kDa) showing that the isolated enzyme contained a few contaminating proteins. The enzyme is a glycoprotein because it binds with high affinity to concanavalin A. It was maximally active at pH 8.0 and its activity was not dependent on bivalent metal ions. The enzyme is a specific phosphatase as it displayed higher affinity for dephosphorylation of large molecular weight phosphate esters. The PPase showed broad substrate specificity for the dephosphorylation of a variety of proteins. The membrane-associated PPase was strongly (70&#8211;80%) inhibited by detergents (0.5%) such as Nonidet P-40, Lubrol PX, Triton X-100 and Tween-20. Pyrophosphate (5 mM) and orthovanadate (400 M) had no significant effect on the activity of the isolated PPase whereas polyamines such as spermine (10 mM) nd spermidine (10 mM) slightly inhibited (20%) the enzymatic activity. Inorganic phosphate (10 mM) and NaF (10 mM), the well-known inhibitors of the cytosolic PPases, had no appreciable effect on the activity of PPase M-I, indicating that the membrane-bound PPase is distinct from the cytosolic PPases. The enzyme was radiolabelled when the intact spermatozoa were subjected to lactoperoxidase-mediated radioiodination reaction. The results show that the PPase M-I is an ecto-enzyme that may play an important role in sperm physiology by causing the dephosphorylation of the sperm outer surface phosphoproteins. 
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