Immunological identification of G protein alpha- and beta-subunits in tail membranes of bovine spermatozoa

Heterotrimeric G proteins are believed to play important roles as signal transducing components in various mammalian sperm functions. To assess the distribution of G proteins in bovine sperm tails, we purified membranes by hypoosmotic swelling of bovine spermatozoa followed by disruption of plasma m...

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Veröffentlicht in:Biology of reproduction 1992-09, Vol.47 (3), p.337-346
Hauptverfasser: Hinsch, K.D. (Universitats-Hautklinik, Giessen, Germany), Hinsch, E, Aumuller, G, Tychowiecka, I, Schultz, G, Schill, W.B
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container_end_page 346
container_issue 3
container_start_page 337
container_title Biology of reproduction
container_volume 47
creator Hinsch, K.D. (Universitats-Hautklinik, Giessen, Germany)
Hinsch, E
Aumuller, G
Tychowiecka, I
Schultz, G
Schill, W.B
description Heterotrimeric G proteins are believed to play important roles as signal transducing components in various mammalian sperm functions. To assess the distribution of G proteins in bovine sperm tails, we purified membranes by hypoosmotic swelling of bovine spermatozoa followed by disruption of plasma membranes in a homogenizer and various centrifugation steps. Electron microscopy revealed highly purified membranes of bovine sperm tails. Subsequently, antisera against synthetic peptides were used to identify G proteins in immunoblots. An antiserum directed against the C-terminal decapeptide of G13 and detecting all known pertussis toxin-sensitive alpha-subunits, reacted specifically with a 40-kDa protein. In contrast, various other specific peptide antisera against alpha-subunits did not detect any G protein in enriched tail membranes. An antiserum recognizing the beta 2-subunit of G proteins and an antiserum reacting with both beta 1- and beta 2-subunits identified a 35-kDa protein in sperm tail membranes. In contrast, antisera against the 36-kDa beta 1-subunit did not detect any relevant proteins in the membrane fraction. Neither G protein alpha-subunits nor G protein beta-subunits were found in the cytosol. Our results suggest that G proteins in membranes of tails of bovine spermatozoa most likely belong to a novel subtype of G protein alpha-subunits, whereas the putative beta-subunit could be identified as a beta 2-subunit
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An antiserum directed against the C-terminal decapeptide of G13 and detecting all known pertussis toxin-sensitive alpha-subunits, reacted specifically with a 40-kDa protein. In contrast, various other specific peptide antisera against alpha-subunits did not detect any G protein in enriched tail membranes. An antiserum recognizing the beta 2-subunit of G proteins and an antiserum reacting with both beta 1- and beta 2-subunits identified a 35-kDa protein in sperm tail membranes. In contrast, antisera against the 36-kDa beta 1-subunit did not detect any relevant proteins in the membrane fraction. Neither G protein alpha-subunits nor G protein beta-subunits were found in the cytosol. 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Psychology</subject><subject>GTP-Binding Proteins - analysis</subject><subject>guanine nucleotide-binding protein</subject><subject>IDENTIFICACION</subject><subject>IDENTIFICATION</subject><subject>Immunoblotting</subject><subject>IMMUNOLOGIE</subject><subject>INMUNOLOGIA</subject><subject>Macromolecular Substances</subject><subject>Male</subject><subject>Mammalian male genital system</subject><subject>Microscopy, Electron</subject><subject>Molecular Sequence Data</subject><subject>Morphology. 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Electron microscopy revealed highly purified membranes of bovine sperm tails. Subsequently, antisera against synthetic peptides were used to identify G proteins in immunoblots. An antiserum directed against the C-terminal decapeptide of G13 and detecting all known pertussis toxin-sensitive alpha-subunits, reacted specifically with a 40-kDa protein. In contrast, various other specific peptide antisera against alpha-subunits did not detect any G protein in enriched tail membranes. An antiserum recognizing the beta 2-subunit of G proteins and an antiserum reacting with both beta 1- and beta 2-subunits identified a 35-kDa protein in sperm tail membranes. In contrast, antisera against the 36-kDa beta 1-subunit did not detect any relevant proteins in the membrane fraction. Neither G protein alpha-subunits nor G protein beta-subunits were found in the cytosol. Our results suggest that G proteins in membranes of tails of bovine spermatozoa most likely belong to a novel subtype of G protein alpha-subunits, whereas the putative beta-subunit could be identified as a beta 2-subunit</abstract><cop>Madison, WI</cop><pub>Society for the Study of Reproduction</pub><pmid>1511086</pmid><doi>10.1095/biolreprod47.3.337</doi><tpages>10</tpages></addata></record>
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ispartof Biology of reproduction, 1992-09, Vol.47 (3), p.337-346
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source MEDLINE; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals
subjects Adenosine Diphosphate Ribose - metabolism
Amino Acid Sequence
Animals
Biological and medical sciences
BOVIDAE
Cattle
Cell Fractionation
Cell Membrane - chemistry
characterization
ESPERMATOZOO
Fundamental and applied biological sciences. Psychology
GTP-Binding Proteins - analysis
guanine nucleotide-binding protein
IDENTIFICACION
IDENTIFICATION
Immunoblotting
IMMUNOLOGIE
INMUNOLOGIA
Macromolecular Substances
Male
Mammalian male genital system
Microscopy, Electron
Molecular Sequence Data
Morphology. Physiology
Pertussis Toxin
PROTEINAS
PROTEINE
Sperm Tail - chemistry
Sperm Tail - ultrastructure
spermatozoa
SPERMATOZOIDE
Vertebrates: reproduction
Virulence Factors, Bordetella - pharmacology
title Immunological identification of G protein alpha- and beta-subunits in tail membranes of bovine spermatozoa
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