Characterization of a low-molecular-mass stimulator protein of Mg2+-independent Ca2+-ATPase: effect on phosphorylation/dephosphorylation, calcium transport and sperm-cell motility.
A 14 kDa cytosolic protein purified from bovine brain homogenate has been recently reported as a stimulator of goat spermatozoa Mg2+-independent Ca2+-ATPase. In the present study, we demonstrate the formation of the [gamma-32P]ATP-labelled phosphoenzyme as the 110 kDa phosphoprotein and its rapid decomposition in presence of the stimulator protein. Together with the cross-reactivity of this 110 kDa protein with an anti-SERCA (sarcoplasmic/endoplasmic reticulum Ca2+-ATPase) 2a antibody, the ATPase can now be conclusively said to belong to the SERCA family, which is activated by the stimulator. The ability of the stimulator to enhance the Ca2+ transport has been elucidated from 45Ca2+ uptake studies and was found to be sensitive to Ca2+ channel blockers. CD revealed an alpha-helical structure of the stimulator. The amino acid analysis suggests that it is composed primarily of hydrophobic and some acidic amino acid residues. The pI of 5.1 has been re-confirmed from two-dimensional electrophoresis. Immuno-cross-reactivity studies indicate that the stimulator or similar proteins are present in cytosolic fractions of liver, kidney or testes in different species, but brain is the richest source. Proteomic analyses of its trypsinized fragments suggest its similarity with bovine THRP (thyroid hormone-responsive protein). The physiological significance of the stimulator has been suggested from its ability to activate sperm-cell motility.
Ghoshal
Srabasti
S
Department of Chemistry, Bose Institute, Kolkata, 700 009, India.
Sengupta
Tanusree
T
Dundung
Sandhya R
SR
Majumder
Gopal C
GC
Sen
Parimal C
PC
D000818:Animals; D001692:Biological Transport; D001921:Brain / Q000378:metabolism; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism*; D002118:Calcium / Q000737:chemistry; D002417:Cattle; D003600:Cytosol / Q000378:metabolism; D006041:Goats; D008274:Magnesium / Q000737:chemistry; D008297:Male; D008861:Microsomes / Q000378:metabolism; D008954:Models, Biological; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism; D010766:Phosphorylation; D013053:Spectrophotometry / Q000379:methods; D013081:Sperm Motility
D010750:Phosphoproteins; D017301:Ca(2+) Mg(2+)-ATPase; D008274:Magnesium; D002118:Calcium