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Molecular Pharmacology
Zaremba, T., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Gierschik, P., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Pines, M., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Bray, P., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Carter, A., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Kahn, R., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Simons, C., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Vinitsky, R., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Goldsmith, P., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Spiegel, A., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Twenty-four of 24 rabbits immunized with the β subunit common to guanine nucleotide binding proteins developed antibodies reactive on immunoblots with the 15-kDa (amino-terminal) tryptic fragment of β. Only 2 of 24 developed antibodies reactive with the 26-kDa (carboxy-terminal) tryptic fragment. The 15-kDa fragment-reactive antibodies were also detected in several nonimmune sera. Antibodies reactive with the 15-kDa fragment could be affinity-purified from all β-immune sera by adsorption to a fusion protein encoded by a cDNA clone identified by expression vector screening. The 15-kDa fragment antibodies in nonimmune sera did not bind to the fusion protein. Limited amino acid sequence homology between the 36-kDa β subunit and the protein encoded by the cDNA clone suggested that the amino-terminal decapeptide of β contains a major epitope. A synthetic decapeptide, corresponding to the amino terminus of the 36-kDa β subunit, effectively and specifically blocked binding of antibodies in β-immune sera (but not in β-reactive nonimmune sera) to nitrocellulose-bound 15-kDa fragment. The 15-kDa fragment-reactive antibodies could be affinity-purified from β-immune sera on a matrix containing bound decapeptide; affinity-purified antibodies reacted equally well with the 36- and 35-kDa forms of the β subunit. Native transducin β/γ complexes readily blocked binding of 15-kDa fragment-reactive antibodies in immune but not nonimmune sera from binding to the nitrocellulose-bound fragment. The results show that nonimmune sera may contain antibodies directed against an epitope of the 15-kDa fragment that is buried in the native β/γ complex. In contrast, the amino terminal decapeptide of the β subunit is exposed on the surface of the native protein and contains a major antigenic site in both the 35- and 36-kDa forms.
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Immunochemical studies of the 36-kDa common β subunit of guanine nucleotide-binding proteins: Identification of a major epitope
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Zaremba, T., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Gierschik, P., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Pines, M., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Bray, P., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Carter, A., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Kahn, R., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Simons, C., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Vinitsky, R., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Goldsmith, P., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Spiegel, A., Molecular Pathophysiology Section, National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD 20892, United States
Immunochemical studies of the 36-kDa common β subunit of guanine nucleotide-binding proteins: Identification of a major epitope
Twenty-four of 24 rabbits immunized with the β subunit common to guanine nucleotide binding proteins developed antibodies reactive on immunoblots with the 15-kDa (amino-terminal) tryptic fragment of β. Only 2 of 24 developed antibodies reactive with the 26-kDa (carboxy-terminal) tryptic fragment. The 15-kDa fragment-reactive antibodies were also detected in several nonimmune sera. Antibodies reactive with the 15-kDa fragment could be affinity-purified from all β-immune sera by adsorption to a fusion protein encoded by a cDNA clone identified by expression vector screening. The 15-kDa fragment antibodies in nonimmune sera did not bind to the fusion protein. Limited amino acid sequence homology between the 36-kDa β subunit and the protein encoded by the cDNA clone suggested that the amino-terminal decapeptide of β contains a major epitope. A synthetic decapeptide, corresponding to the amino terminus of the 36-kDa β subunit, effectively and specifically blocked binding of antibodies in β-immune sera (but not in β-reactive nonimmune sera) to nitrocellulose-bound 15-kDa fragment. The 15-kDa fragment-reactive antibodies could be affinity-purified from β-immune sera on a matrix containing bound decapeptide; affinity-purified antibodies reacted equally well with the 36- and 35-kDa forms of the β subunit. Native transducin β/γ complexes readily blocked binding of 15-kDa fragment-reactive antibodies in immune but not nonimmune sera from binding to the nitrocellulose-bound fragment. The results show that nonimmune sera may contain antibodies directed against an epitope of the 15-kDa fragment that is buried in the native β/γ complex. In contrast, the amino terminal decapeptide of the β subunit is exposed on the surface of the native protein and contains a major antigenic site in both the 35- and 36-kDa forms.
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