ChemBioChem 2011-03-07

The chemoselective one-step alkylation and isolation of thiophosphorylated cdk2 substrates in the presence of native cysteine.

Sarah E Lee, Lucy M Elphick, Holger B Kramer, Alexandra M E Jones, Emma S Child, Alexandra A Anderson, Laurent Bonnac, Natsuko Suwaki, Benedikt M Kessler, Véronique Gouverneur, David J Mann

Index: ChemBioChem. 12(4) , 633-40, (2011)

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Abstract

The elucidation of signalling pathways relies heavily upon the identification of protein kinase substrates. Recent investigations have demonstrated the efficacy of chemical genetics using ATP analogues and modified protein kinases for specific substrate labelling. Here we combine N(6) -(cyclohexyl)ATPγS with an analogue-sensitive cdk2 variant to thiophosphorylate its substrates and demonstrate a pH-dependent, chemoselective, one-step alkylation to facilitate the detection or isolation of thiophosphorylated peptides.Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.