IMR Press / FBS / Volume 5 / Issue 2 / DOI: 10.2741/S397

Frontiers in Bioscience-Scholar (FBS) is published by IMR Press from Volume 13 Issue 1 (2021). Previous articles were published by another publisher on a subscription basis, and they are hosted by IMR Press on as a courtesy and upon agreement with Frontiers in Bioscience.

Catalytic site amino acids of PKGI-alpha influence allosteric cGMP binding
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1 Department of Biology, Wheaton College, Wheaton, IL 60187
2 Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN 37232-0615
3 Department of Biochemistry, Vanderbilt University, Nashville, TN 37232-0615

*Author to whom correspondence should be addressed.

Front. Biosci. (Schol Ed) 2013, 5(2), 650–660;
Published: 1 January 2013

Ser-64, an autophosphorylation site in the autoinhibitory subdomain of cGMP-dependent protein kinase type I-alpha (PKGI-alpha), lowers affinity for cGMP and suppresses catalytic activity (1). Using the structure of homologous cAMP-dependent protein kinase as a model, three conserved residues (Gln-401, His-404, Cys-518) in the PKGI-alpha catalytic site are predicted to be juxtaposed to Ser-64 (2). Individual point mutants (Q401A, H404A and C518A) and a double mutant (S64A/H404A) have been generated. cGMP or cAMP affinities (Ka) of each mutant protein for phosphotransferase activation and allosteric [3H]cGMP-binding affinity (KD) of each mutant protein are significantly improved over those of wild-type (WT) PKGI-alpha. However, affinities (Km) of the mutant PKGs for peptide substrates or ATP are unaltered. Kinase activity ratio (-GMP/+cGMP) of H404A is greater than that for WT, Q401A, or C518A, and similar to that for S64A and S64A/H404A. These results reveal a unique mechanism whereby catalytic domain residues predicted to be spatially close to Ser-64 of the regulatory domain weaken the intrinsically high affinity of PKGI-alpha for cGMP and provide for autoinhibition of catalytic activity.

cGMP-dependent protein kinase
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