Linked Life Data

16128581

Source:http://linkedlifedata.com/resource/pubmed/id/16128581

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
35
pubmed:dateCreated
2005-8-30
pubmed:abstractText
The yeast Saccharomyces cerevisiae alpha-factor pheromone receptor (Ste2p) was used as a model G protein-coupled receptor (GPCR). A 73-mer multidomain fragment of Ste2p (residues 267-339) containing the third extracellular loop, the seventh transmembrane domain, and 40 residues of the cytosolic tail (E3-M7-24-T40) was biosynthesized fused to a carrier protein. The multidomain fusion protein (designated M7FP) was purified to near homogeneity as judged by HPLC and characterized by mass spectrometry. In minimal medium, 30-40 mg of M7FP were obtained per liter of culture. The 73-residue peptide was released from its carrier by CNBr and obtained in wild-type, (15)N, and (13)C/(15)N forms. The E3-M7-24-T40 peptide integrated into 1-palmitoyl-2-hydroxy-sn-glycero-3-[phospho-rac-(1-glycerol)] and dodecylphosphocholine micelles at concentrations (200-500 microM) suitable for NMR investigations. HSQC experiments performed in organic solvents and detergent micelles on (15)N-labeled E3-M7-24-T40 showed a clear dispersion of the nitrogen-amide proton correlation cross-peaks indicative of a pure, uniformly labeled molecule that assumed a partially ordered structure. NOE connectivities, chemical shift indices, J-coupling analysis, and structural modeling suggested that in trifluoroethanol/water (1:1) helical subdomains existed in both the transmembrane and cytoslic tail of the multidomain peptide. Similar conclusions were reached in chloroform/methanol/water (4:4:1). As the cytosolic tail participates in down-regulation of Ste2p, the helical regions in the Ste2p tail may play a role in protein-protein interactions involved in endocytosis.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Sep
pubmed:issn
0006-2960
pubmed:author
pubmed:issnType
Print
pubmed:day
6
pubmed:volume
44
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
11795-810
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed-meshheading:16128581-Amino Acid Sequence, pubmed-meshheading:16128581-Chromatography, High Pressure Liquid, pubmed-meshheading:16128581-Cyanogen Bromide, pubmed-meshheading:16128581-Mass Spectrometry, pubmed-meshheading:16128581-Molecular Sequence Data, pubmed-meshheading:16128581-Nitrogen Isotopes, pubmed-meshheading:16128581-Nuclear Magnetic Resonance, Biomolecular, pubmed-meshheading:16128581-Protein Conformation, pubmed-meshheading:16128581-Protein Structure, Secondary, pubmed-meshheading:16128581-Protein Structure, Tertiary, pubmed-meshheading:16128581-Receptors, G-Protein-Coupled, pubmed-meshheading:16128581-Receptors, Mating Factor, pubmed-meshheading:16128581-Recombinant Fusion Proteins, pubmed-meshheading:16128581-Saccharomyces cerevisiae, pubmed-meshheading:16128581-Saccharomyces cerevisiae Proteins
pubmed:year
2005
pubmed:articleTitle
Biosynthesis and NMR analysis of a 73-residue domain of a Saccharomyces cerevisiae G protein-coupled receptor.
pubmed:affiliation
Department of Chemistry, College of Staten Island and Macromolecular Assemblies Institute of the City University of New York, Staten Island, New York 10314, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, U.S. Gov't, Non-P.H.S., Research Support, N.I.H., Extramural