MULTIPLE GTP-BINDING PROTEINS IN SEA-URCHIN SPERM - EVIDENCE FOR GS AND SMALL G-PROTEINS

被引:0
作者
CUELLARMATA, P
MARTINEZCADENA, G
CASTELLANO, LE
ALDANAVELOZ, G
NOVOAMARTINEZ, G
VARGAS, I
DARSZON, A
GARCIASOTO, J
机构
[1] UNIV GUANAJUATO, FAC QUIM, INST INVEST BIOL EXPTL, GUANAJUATO 36000, MEXICO
[2] IPN, CINVESTAV, DEPT BIOQUIM, CUERNAVACA, MORELOS, MEXICO
[3] Univ Nacl Autonoma Mexico, INST BIOTECNOL, CUERNAVACA, MORELOS, MEXICO
关键词
ADP-RIBOSYLATION; CHOLERA TOXIN; G-PROTEIN; RAS; SEA URCHIN SPERM;
D O I
暂无
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
Sea urchin sperm plasma membranes isolated from heads and flagella were used to examine the presence of Gs (stimulatory guanine nucleotide-binding regulatory protein) and small G-proteins. Flagellar plasma membranes incubated with [P-32]NAD and cholera toxin (CTX) displayed radiolabeling in a protein of 48 kDa, which was reactive by immunoblotting with a specific antibody against mammalian Gs. CTX-catalyzed [P-32]ADP-ribosylation in conjunction with immunoprecipitation with anti-Gs, followed by electrophoresis and autoradiography, revealed one band of 48 kDa. Head plasma membranes, in contrast, did not show substrates for ADP-ribosylation by CTX. In flagellar and head plasma membranes pertussis toxin (PTX) ADP-ribosylated the same protein described previously in membranes from whole sperm; the extent of ADP-ribosylation by PTX was higher in flagellar than in head membranes. Small G-proteins were investigated by [P-32]GTP-blotting. Both head and flagellar plasma membranes showed three radiolabeled bands of 28, 25 and 24 kDa. Unlabeled GTP and GDP, but not other nucleotides, interfered with the [alpha-P-32]GTP-binding in a concentration-dependent manner. A monoclonal antibody against human Ras p21 recognized a single protein of 21 kDa only in flagellar membranes. Thus, sea urchin sperm contain a membrane protein that shares characteristics with mammalian Gs and four small G-proteins, including Res. Gs, Gi and Ras are enriched in flagellar membranes while the other small G-proteins do not display a preferential distribution along the sea urchin sperm plasma membrane. The role of these G-proteins in sea urchin sperm is presently under investigation.
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页码:173 / 181
页数:9
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