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PMID: 16981699 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S.

Minimal determinants for binding activated G alpha from the structure of a G alpha(i1)-peptide dimer.

Biochemistry ·Vol. 45 ·No. 38 ·2006-09-26 ·Pages 11390-400

Johnston CA, Lobanova ES, Shavkunov AS, Low J, Ramer JK, Blaesius R, Fredericks Z, Willard FS, Kuhlman B, Arshavsky VY, Siderovski DP

Abstract

G-proteins cycle between an inactive GDP-bound state and an active GTP-bound state, serving as molecular switches that coordinate cellular signaling. We recently used phage display to identify a series of peptides that bind G alpha subunits in a nucleotide-dependent manner [Johnston, C. A., Willard, F. S., Jezyk, M. R., Fredericks, Z., Bodor, E. T., Jones, M. B., Blaesius, R., Watts, V. J., Harden, T. K., Sondek, J., Ramer, J. K., and Siderovski, D. P. (2005) Structure 13, 1069-1080]. Here we describe the structural features and functions of KB-1753, a peptide that binds selectively to GDP x AlF4(-)- and GTPgammaS-bound states of G alpha(i) subunits. KB-1753 blocks interaction of G alpha(transducin) with its effector, cGMP phosphodiesterase, and inhibits transducin-mediated activation of cGMP degradation. Additionally, KB-1753 interferes with RGS protein binding and resultant GAP activity. A fluorescent KB-1753 variant was found to act as a sensor for activated G alpha in vitro. The crystal structure of KB-1753 bound to G alpha(i1) x GDP x AlF4(-) reveals binding to a conserved hydrophobic groove between switch II and alpha3 helices and, along with supporting biochemical data and previous structural analyses, supports the notion that this is the site of effector interactions for G alpha(i) subunits.

MeSH Terms
Aluminum Compounds/chemistry Amino Acid Sequence Amino Acid Substitution Bacterial Proteins/metabolism Crystallography, X-Ray Dimerization Fluorides/chemistry GTP-Binding Protein alpha Subunits, Gi-Go/chemistry,metabolism Guanosine Diphosphate/chemistry Humans Luminescent Proteins/metabolism Models, Molecular Molecular Sequence Data Protein Binding Protein Structure, Secondary RGS Proteins/metabolism Recombinant Fusion Proteins/metabolism Structure-Activity Relationship
Chemicals
Aluminum Compounds Bacterial Proteins Luminescent Proteins RGS Proteins Recombinant Fusion Proteins yellow fluorescent protein, Bacteria Guanosine Diphosphate tetrafluoroaluminate GTP-Binding Protein alpha Subunits, Gi-Go Fluorides
Authors & Affiliations
11 authors, click to expand affiliations / ORCID
Johnston Christopher A
Department of Pharmacology, University of North Carolina School of Medicine, Chapel Hill, North Carolina 27599-7365, USA.
Lobanova Ekaterina S
Shavkunov Alexander S
Low Justin
Ramer J Kevin
Blaesius Rainer
Fredericks Zoey
Willard Francis S
Kuhlman Brian
Arshavsky Vadim Y
Siderovski David P
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Article Info
Journal
Biochemistry
Abbr.
Biochemistry
ISSN
0006-2960
DOI
10.1021/bi0613832
Published
2006-09-26
Pages
11390-400
Language
English
Region
United States
NLM ID
0370623
PMCID
PMC2597383
Subset
IM
Grants
NIGMS NIH HHS · R01 GM074268-01A1 · United States
NIGMS NIH HHS · F32 GM076944-01 · United States
NIGMS NIH HHS · R01 GM074268-02 · United States
NEI NIH HHS · EY12859 · United States
NIGMS NIH HHS · R01 GM074268-03 · United States
NIGMS NIH HHS · 1 F32 GM076944 · United States
NIGMS NIH HHS · R01 GM074268 · United States
NIGMS NIH HHS · F32 GM076944-02 · United States
NEI NIH HHS · R01 EY012859 · United States
NEI NIH HHS · R01 EY012859-06 · United States
NIGMS NIH HHS · F32 GM076944 · United States
Databases
PDB
2G83
External Links
PubMed source DOI: 10.1021/bi0613832 PMC Full Text View journal details
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