Document Detail


A predicted amphipathic helix mediates plasma membrane localization of GRK5.
MedLine Citation:
PMID:  14976207     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
G protein-coupled receptor kinases (GRKs) specifically phosphorylate agonist-occupied G protein-coupled receptors at the inner surface of the plasma membrane (PM), leading to receptor desensitization. GRKs utilize a variety of mechanisms to bind tightly, and sometimes reversibly, to cellular membranes. Previous studies demonstrated the presence of a membrane binding domain in the C terminus of GRK5. Here we define a mechanism by which this short C-terminal stretch of amino acids of GRK5 mediates PM localization. Secondary structure predictions suggest that a region contained within amino acids 546-565 of GRK5 forms an amphipathic helix, with the key features of the predicted helix being a hydrophobic patch of amino acids on one face of the helix, hydrophilic amino acids on the opposite face, and a number of basic amino acids surrounding the hydrophobic patch. We show that amino acids 546-565 of GRK5 are sufficient to target the cytoplasmic green fluorescent protein (GFP) to the PM, and the hydrophobic amino acids are necessary for PM targeting of GFP-546-565. Moreover, full-length GRK5-GFP is localized to the PM, but mutation of the hydrophobic patch or the surrounding basic amino acids prevents PM localization of GRK5-GFP. Last, we show that mutation of the hydrophobic residues severely diminishes phospholipid-dependent autophosphorylation of GRK5 and phosphorylation of membrane-bound rhodopsin by GRK5. The findings in this report thus suggest the presence of a membrane binding motif in GRK5 and define the importance of a group of hydrophobic amino acids within this motif in mediating its PM localization.
Authors:
Manimekalai M Thiyagarajan; RoseAnn P Stracquatanio; Alexey N Pronin; Daniel S Evanko; Jeffrey L Benovic; Philip B Wedegaertner
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, P.H.S.     Date:  2004-02-19
Journal Detail:
Title:  The Journal of biological chemistry     Volume:  279     ISSN:  0021-9258     ISO Abbreviation:  J. Biol. Chem.     Publication Date:  2004 Apr 
Date Detail:
Created Date:  2004-04-19     Completed Date:  2004-06-10     Revised Date:  2007-11-15    
Medline Journal Info:
Nlm Unique ID:  2985121R     Medline TA:  J Biol Chem     Country:  United States    
Other Details:
Languages:  eng     Pagination:  17989-95     Citation Subset:  IM    
Affiliation:
Department of Microbiology and Immunology and Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
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MeSH Terms
Descriptor/Qualifier:
Amino Acid Motifs
Amino Acid Sequence
Animals
COS Cells
Cell Line
Cell Membrane / metabolism
G-Protein-Coupled Receptor Kinase 5
Green Fluorescent Proteins
Humans
Luminescent Proteins / metabolism
Microscopy, Confocal
Molecular Sequence Data
Mutation
Phospholipids / chemistry
Phosphorylation
Plasmids / metabolism
Protein Binding
Protein Structure, Secondary
Protein Structure, Tertiary
Protein-Serine-Threonine Kinases / biosynthesis*,  chemistry
Rhodopsin / chemistry
Sequence Homology, Amino Acid
Time Factors
Tubulin / chemistry
Grant Support
ID/Acronym/Agency:
GM44944/GM/NIGMS NIH HHS; GM56444/GM/NIGMS NIH HHS; GM62884/GM/NIGMS NIH HHS
Chemical
Reg. No./Substance:
0/Luminescent Proteins; 0/Phospholipids; 0/Tubulin; 147336-22-9/Green Fluorescent Proteins; 9009-81-8/Rhodopsin; EC 2.7.11.1/Protein-Serine-Threonine Kinases; EC 2.7.11.16/G-Protein-Coupled Receptor Kinase 5; EC 2.7.11.16/GRK5 protein, human

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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