The protein inhibitor of adenosine 3′,5′-monophosphate-dependent protein kinases the NH2-terminal portion of the peptide chain contains the inhibitory site |
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Authors: | Jacques G Demaille Concepción Ferraz Edmond H Fischer |
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Institution: | 1. Centre de Recherches de Biochimie Macromoléculaire, CNRS, BP 5051, F-34033 Montpellier Cedex France;2. Department of Biochemistry SJ-70, University of Washington, Seatlle, WA 98195 U.S.A. |
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Abstract: | The protein inhibitor of adenosine 3′,5′-monophosphate-dependent protein kinases from skeletal muscle was subjected to various chemical and enzymatic treatments in an attempt to delineate the part of the molecule responsible for the interaction with the catalytic subunit of the kinase. Only a small portion of the chain seems to be required since thermolysin and staphylococcal protease digestions do not abolish the inhibitory properties. This inhibitory site must contain the essential arginyl side chain(s), whereas lysyl and carboxylic side chains do not appear to be involved in the interaction with the catalytic subunit.Digestion of the COOH-terminus of the inhibitor by carboxypeptidase Y results in a doubling of the Ki value. On the other hand, an inhibitory pentadecapeptide (Ki = 25 nM), presumably NH2-terminal in the entire molecule, could be isolated from a staphylococcal protease digest by means of gel filtration followed by ion exchange on phosphocellulose. The purified inhibitory peptide contains two out of the four arginyl residues present in the entire molecule. The remarkable affinity and specificity of the protein kinase inhibitor for the catalytic subunit of adenosine 3′,5′-monophosphate-dependent protein kinases may thus be tentatively explained on the basic of a two-prong attachment of the inhibitor. The NH2-terminal portion of the chain would bind at the substate binding site, whereas the COOH-terminal part would be held elsewhere. |
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Keywords: | Protein kinase inhibitor Cyclic nucleotide Hormone action Peptide inhibitor Inhibitory site C the catalytic subunits of the cyclic AMP-dependent protein kinase R the regulatory subunits of the cyclic AMP-dependent protein kinase |
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