Work

Dissecting the Cooperative Reassociation of the Regulatory and Catalytic Subunits of cAMP-dependent Protein Kinase

Year: 1997

Type: article

Abstract: The catalytic (C) subunit of cAMP-dependent protein kinase requires two distinct surfaces to form a stable complex with its physiological inhibitors, the regulatory (R) subunits and the heat-stable protein kinase inhibitors. In addition to a substrate-like segment that is common to both inhibitors, R requires a peripheral recognition site, PRS2. This surface is comprised of the essential phosphorylation site, Thr-197, His-87, Trp-196, and several surrounding basic residues. To probe the role of Trp-196 in the recognition of R, Trp-196 was replaced with Arg and Ala. Although both rC(W196A) and rC(W196R) were inhibited readily with cAMP-free R, they failed to form an inhibited holoenzyme complex with native R under conditions in which wild-type holoenzyme formed readily. Pairing rC(W196R) with mutant forms of R lacking domain B or having defects in cAMP binding sites A or B highlighted the importance of the conformation of R, and, in particular, the accessibility of site A. One of these mutants, rR(R333K), having a defect in cAMP binding site B formed a stable complex with rC(W196R) in the absence of cAMP. However, unlike wild-type holoenzyme, this complex was active.

Source: Journal of Biological Chemistry

DOI: 10.1074/jbc.272.51.31998

Authors: Robin M. Gibson, Susan S. Taylor

Institutions: Howard Hughes Medical Institute, University of California San Diego

Language: English

Topic: Protein Kinase Regulation and GTPase Signaling

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: hybrid

Estimated APC paid: $3,430

Funders: National Cancer Institute, National Institute of General Medical Sciences