Slow phosphoryl transfer of the gamma phosphate from the ATP analog AMP-PNP onto a synthetic substrate peptide, PKS, was captured in crystals of protein kinase A, according to a recent report.1  PKS, a twenty residue peptide, is identical to residues 5-24 of the endogenous inhibitor peptide PKI (PKI5-24), except that N20 is replaced by alanine and A21 is replaced by serine.  While synthesis of isotopically-labeled PKS for NMR studies would be prohibitively expensive, a method was developed previously for expressing and purifying isotopically-enriched substrate and inhibitor peptides using a recombinant fusion protein system.2  The purpose of this study is to express and purify PKS as a recombinant fusion protein to support NMR studies of protein kinase A in complex with PKS. 1Bastidas, A.C.; Deal, M.S.; Steichen, J.M.; Yurong, G.; Wu, J.; Taylor, S.S. J. Am. Chem. Soc. 2013 , 135, 4788-4798.2Masterson, L.R.; Bortone, N.; Yu, T.; Ha, K.M.; Gaffarogullari, E.C.; Nguyen, O.; Veglia, G.  Protein Expression and Purification 2009 , 64, 231-236.

Production of PKS for NMR studies of cAMP dependent protein kinase A

247th National Meeting (2014)

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