Antisera against a guanine nucleotide binding protein from retina cross-react with the β subunit of the adenyl cyclase-associated guanine nucleotide binding proteins, N(s) and N(i)

P. Gierschik, J. Codina, C. Simons, L. Birnbaumer, Allen M. Spiegel

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Abstract

Antisera were produced in rabbits against a guanine nucleotide binding protein (N protein), transducin, purified from bovine retina. Antiserum AS/1, which recognized all three subunits (α, β, and γ) of the holoprotein, was tested for cross-reactivity with the subunits of the adenylyl cyclase [adenylate cyclase; ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1]-associated stimulatory (N(s)) and inhibitory (N(i)) N proteins purified from human erythrocytes. AS/1 showed strong reactivity against the β subunits of both N(s) and N(i) but failed to cross-react with either the α or γ subunits of N(s) and N(i). Seven additional antisera against transducin reacted with the β subunits but not with the α or γ subunits of N(s) and N(i). A single antiserum against transducin reacted with the α subunit of N(i) but not of N(s). Immunostaining of the β subunits of N(s) and N(i) was proportional to the amount of β subunit blotted and to the antiserum concentration. Immunostaining of either human erythrocyte or bovine cerebral cortical plasma membrane proteins with AS/1 showed a single band, comigrating with the β subunit of transducin; this band was absent in bovine erythrocyte membranes. Estimation of the amount of β subunit by immunoblotting with AS/1 showed that the β subunit comprises ≃ 2% of bovine cerebral cortical plasma membrane protein, ≃ 100-fold more than in human erythrocyte membranes. These findings provide immunochemical evidence for similarities in the β subunits and differences in the α and γ subunits of this family of N proteins. Antisera against transducin react specifically with the β subunits of N(s) and N(i) in crude plasma membranes and, thus, can serve as specific probes for the β subunit.

Original languageEnglish (US)
Pages (from-to)727-731
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume82
Issue number3
DOIs
StatePublished - 1985
Externally publishedYes

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Guanine Nucleotides
Transducin
Adenylyl Cyclases
Retina
Immune Sera
Carrier Proteins
Cell Membrane
Erythrocyte Membrane
Blood Proteins
Membrane Proteins
Erythrocytes
Immunoblotting
Proteins
Rabbits

ASJC Scopus subject areas

  • Genetics
  • General

Cite this

@article{582df349e99b4958b1446bde31919811,
title = "Antisera against a guanine nucleotide binding protein from retina cross-react with the β subunit of the adenyl cyclase-associated guanine nucleotide binding proteins, N(s) and N(i)",
abstract = "Antisera were produced in rabbits against a guanine nucleotide binding protein (N protein), transducin, purified from bovine retina. Antiserum AS/1, which recognized all three subunits (α, β, and γ) of the holoprotein, was tested for cross-reactivity with the subunits of the adenylyl cyclase [adenylate cyclase; ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1]-associated stimulatory (N(s)) and inhibitory (N(i)) N proteins purified from human erythrocytes. AS/1 showed strong reactivity against the β subunits of both N(s) and N(i) but failed to cross-react with either the α or γ subunits of N(s) and N(i). Seven additional antisera against transducin reacted with the β subunits but not with the α or γ subunits of N(s) and N(i). A single antiserum against transducin reacted with the α subunit of N(i) but not of N(s). Immunostaining of the β subunits of N(s) and N(i) was proportional to the amount of β subunit blotted and to the antiserum concentration. Immunostaining of either human erythrocyte or bovine cerebral cortical plasma membrane proteins with AS/1 showed a single band, comigrating with the β subunit of transducin; this band was absent in bovine erythrocyte membranes. Estimation of the amount of β subunit by immunoblotting with AS/1 showed that the β subunit comprises ≃ 2{\%} of bovine cerebral cortical plasma membrane protein, ≃ 100-fold more than in human erythrocyte membranes. These findings provide immunochemical evidence for similarities in the β subunits and differences in the α and γ subunits of this family of N proteins. Antisera against transducin react specifically with the β subunits of N(s) and N(i) in crude plasma membranes and, thus, can serve as specific probes for the β subunit.",
author = "P. Gierschik and J. Codina and C. Simons and L. Birnbaumer and Spiegel, {Allen M.}",
year = "1985",
doi = "10.1073/pnas.82.3.727",
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T1 - Antisera against a guanine nucleotide binding protein from retina cross-react with the β subunit of the adenyl cyclase-associated guanine nucleotide binding proteins, N(s) and N(i)

AU - Gierschik, P.

AU - Codina, J.

AU - Simons, C.

AU - Birnbaumer, L.

AU - Spiegel, Allen M.

PY - 1985

Y1 - 1985

N2 - Antisera were produced in rabbits against a guanine nucleotide binding protein (N protein), transducin, purified from bovine retina. Antiserum AS/1, which recognized all three subunits (α, β, and γ) of the holoprotein, was tested for cross-reactivity with the subunits of the adenylyl cyclase [adenylate cyclase; ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1]-associated stimulatory (N(s)) and inhibitory (N(i)) N proteins purified from human erythrocytes. AS/1 showed strong reactivity against the β subunits of both N(s) and N(i) but failed to cross-react with either the α or γ subunits of N(s) and N(i). Seven additional antisera against transducin reacted with the β subunits but not with the α or γ subunits of N(s) and N(i). A single antiserum against transducin reacted with the α subunit of N(i) but not of N(s). Immunostaining of the β subunits of N(s) and N(i) was proportional to the amount of β subunit blotted and to the antiserum concentration. Immunostaining of either human erythrocyte or bovine cerebral cortical plasma membrane proteins with AS/1 showed a single band, comigrating with the β subunit of transducin; this band was absent in bovine erythrocyte membranes. Estimation of the amount of β subunit by immunoblotting with AS/1 showed that the β subunit comprises ≃ 2% of bovine cerebral cortical plasma membrane protein, ≃ 100-fold more than in human erythrocyte membranes. These findings provide immunochemical evidence for similarities in the β subunits and differences in the α and γ subunits of this family of N proteins. Antisera against transducin react specifically with the β subunits of N(s) and N(i) in crude plasma membranes and, thus, can serve as specific probes for the β subunit.

AB - Antisera were produced in rabbits against a guanine nucleotide binding protein (N protein), transducin, purified from bovine retina. Antiserum AS/1, which recognized all three subunits (α, β, and γ) of the holoprotein, was tested for cross-reactivity with the subunits of the adenylyl cyclase [adenylate cyclase; ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1]-associated stimulatory (N(s)) and inhibitory (N(i)) N proteins purified from human erythrocytes. AS/1 showed strong reactivity against the β subunits of both N(s) and N(i) but failed to cross-react with either the α or γ subunits of N(s) and N(i). Seven additional antisera against transducin reacted with the β subunits but not with the α or γ subunits of N(s) and N(i). A single antiserum against transducin reacted with the α subunit of N(i) but not of N(s). Immunostaining of the β subunits of N(s) and N(i) was proportional to the amount of β subunit blotted and to the antiserum concentration. Immunostaining of either human erythrocyte or bovine cerebral cortical plasma membrane proteins with AS/1 showed a single band, comigrating with the β subunit of transducin; this band was absent in bovine erythrocyte membranes. Estimation of the amount of β subunit by immunoblotting with AS/1 showed that the β subunit comprises ≃ 2% of bovine cerebral cortical plasma membrane protein, ≃ 100-fold more than in human erythrocyte membranes. These findings provide immunochemical evidence for similarities in the β subunits and differences in the α and γ subunits of this family of N proteins. Antisera against transducin react specifically with the β subunits of N(s) and N(i) in crude plasma membranes and, thus, can serve as specific probes for the β subunit.

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U2 - 10.1073/pnas.82.3.727

DO - 10.1073/pnas.82.3.727

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JF - Proceedings of the National Academy of Sciences of the United States of America

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