Affinity methods for phosphorylation-dependent interactions.

TY - JOUR

T1 - Affinity methods for phosphorylation-dependent interactions.

AU - Moorhead, Greg

AU - MacKintosh, Carol

PY - 2004/1/1

Y1 - 2004/1/1

N2 - 14-3-3s are a highly conserved protein family that exert many regulatory roles in eukaryotic cells by binding to phosphopeptide motifs in diverse target proteins. Here, we describe 14-3-3 affinity binding procedures that can be used to purify and identify 14-3-3-binding phosphoproteins; monitor how their phosphorylation and 14-3-3 binding is regulated by extracellular stimuli; define the functional effects of 14-3-3s on individual targets; and identify relevant protein phosphatases and kinases. In principle, these methods could be adapted to characterize other types of protein-protein interaction that depend on covalent modification of target sites.

AB - 14-3-3s are a highly conserved protein family that exert many regulatory roles in eukaryotic cells by binding to phosphopeptide motifs in diverse target proteins. Here, we describe 14-3-3 affinity binding procedures that can be used to purify and identify 14-3-3-binding phosphoproteins; monitor how their phosphorylation and 14-3-3 binding is regulated by extracellular stimuli; define the functional effects of 14-3-3s on individual targets; and identify relevant protein phosphatases and kinases. In principle, these methods could be adapted to characterize other types of protein-protein interaction that depend on covalent modification of target sites.

KW - Protein-protein interactions

KW - affinity chromatography

KW - phosphorylation

KW - phosphopeptide

UR - https://www.scopus.com/pages/publications/3242664590

U2 - 10.1385/1-59259-762-9:469

DO - 10.1385/1-59259-762-9:469

M3 - Review article

C2 - 15064476

AN - SCOPUS:3242664590

SN - 1064-3745

VL - 261

SP - 469

EP - 478

JO - Methods in Molecular Biology (Clifton, N.J.)

JF - Methods in Molecular Biology (Clifton, N.J.)

ER -