Title: Substrate-Specific Reorganization of the Conformational Ensemble of CSK Implicates Novel Modes of Kinase Function

Abstract

Protein kinases use ATP as a phosphoryl donor for the posttranslational modification of signaling targets. It is generally thought that the binding of this nucleotide induces conformational changes leading to closed, more compact forms of the kinase domain that ideally orient active-site residues for efficient catalysis. The kinase domain is oftentimes flanked by additional ligand binding domains that up- or down-regulate catalytic function. C-terminal Src kinase (Csk) is a multidomain tyrosine kinase that is up-regulated by N-terminal SH2 and SH3 domains. Although the X-ray structure of Csk suggests the enzyme is compact, X-ray scattering studies indicate that the enzyme possesses both compact and open conformational forms in solution. Here, we investigated whether interactions with the ATP analog AMP-PNP and ADP can shift the conformational ensemble of Csk in solution using a combination of small angle x-ray scattering and molecular dynamics simulations. We find that binding of AMP-PNP shifts the ensemble towards more extended rather than more compact conformations. Binding of ADP further shifts the ensemble towards extended conformations, including highly extended conformations not adopted by the apo protein, nor by the AMP-PNP bound protein. These ensembles indicate that any compaction of the kinase domain induced by nucleotide binding does notmore » extend to the overall multi-domain architecture. Instead, assembly of an ATP-bound kinase domain generates further extended forms of Csk that may have relevance for kinase scaffolding and Src regulation in the cell.« less

Authors:

Jamros, Michael A. ^[1]; Oliveira, Leandro C. ^[2]; Whitford, Paul C. ^[3]; Onuchic, José N. ^[3]; Adams, Joseph A. ^[4]; Jennings, Patricia A. ^[1]

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+ Show Author Affiliations

1. Univ. of California, San Diego, La Jolla, CA (United States). Dept. of Chemistry and Biochemistry
2. Laboratorio Nacional de Ciencia e Tecnologia do Bioetanol – CTBE/CNPEM, Campinas, Sao Paulo (Brazil)
3. Rice Univ., Houston, TX (United States). Center for Theoretical Biological Physics
4. Univ. of California, San Diego, La Jolla, CA (United States). Dept. of Pharmacology

Publication Date:

Thu Sep 20 00:00:00 EDT 2012

Research Org.:

SLAC National Accelerator Laboratory (SLAC), Menlo Park, CA (United States). Stanford Synchrotron Radiation Lightsource (SSRL)

Sponsoring Org.:

USDOE Office of Science (SC), Biological and Environmental Research (BER); USDOE Office of Science (SC), Basic Energy Sciences (BES)

OSTI Identifier:

1627225

Grant/Contract Number:  

AC02-76SF00515

Resource Type:

Accepted Manuscript

Journal Name:

PLoS Computational Biology (Online)

Additional Journal Information:

Journal Name: PLoS Computational Biology (Online); Journal Volume: 8; Journal Issue: 9; Journal ID: ISSN 1553-7358

Publisher:

Public Library of Science

Country of Publication:

United States

Language:

English

Subject:

59 BASIC BIOLOGICAL SCIENCES; Biochemistry & Molecular Biology; Mathematical & Computational Biology; Nucleotides; Small-angle scattering; Membrane proteins; Protein kinases; SH2 domains; Curve fitting; Molecular dynamics; Phosphorylation