Linked Life Data

15481977

Source:http://linkedlifedata.com/resource/pubmed/id/15481977

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PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
22
pubmed:dateCreated
2004-10-14
pubmed:abstractText
Human P2Y receptors encompass at least eight subtypes of Class A G protein-coupled receptors (GPCRs), responding to adenine and/or uracil nucleotides. Using a BLAST search against the Homo sapiens subset of the SWISS-PROT and TrEMBL databases, we identified 68 proteins showing high similarity to P2Y receptors. To address the problem of low sequence identity between rhodopsin and the P2Y receptors, we performed a multiple-sequence alignment of the retrieved proteins and the template bovine rhodopsin, combining manual identification of the transmembrane domains (TMs) with automatic techniques. The resulting phylogenetic tree delineated two distinct subgroups of P2Y receptors: Gq-coupled subtypes (e.g., P2Y1) and those coupled to Gi (e.g., P2Y12). On the basis of sequence comparison we mutated three Tyr residues of the putative P2Y1 binding pocket to Ala and Phe and characterized pharmacologically the mutant receptors expressed in COS-7 cells. The mutation of Y306 (7.35, site of a cationic residue in P2Y12) or Y203 in the second extracellular loop selectively decreased the affinity of the agonist 2-MeSADP, and the Y306F mutation also reduced antagonist (MRS2179) affinity by 5-fold. The Y273A (6.48) mutation precluded the receptor activation without a major effect on the ligand-binding affinities, but the Y273F mutant receptor still activated G proteins with full agonist affinity. Thus, we have identified new recognition elements to further define the P2Y1 binding site and related these to other P2Y receptor subtypes. Following sequence-based secondary-structure prediction, we constructed complete models of all the human P2Y receptors by homology to rhodopsin. Ligand docking on P2Y1 and P2Y12 receptor models was guided by mutagenesis results, to identify the residues implicated in the binding process. Different sets of cationic residues in the two subgroups appeared to coordinate phosphate-bearing ligands. Within the P2Y1 subgroup these residues are R3.29, K/R6.55, and R7.39. Within the P2Y12 subgroup, the only residue in common with P2Y1 is R6.55, and the role of R3.29 in TM3 seems to be fulfilled by a Lys residue in EL2, whereas the R7.39 in TM7 seems to be substituted by K7.35. Thus, we have identified common and distinguishing features of P2Y receptor structure and have proposed modes of ligand binding for the two representative subtypes that already have well-developed ligands.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
0022-2623
pubmed:author
pubmed:issnType
Print
pubmed:day
21
pubmed:volume
47
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
5393-404
pubmed:dateRevised
2010-11-18
pubmed:meshHeading
pubmed-meshheading:15481977-Amino Acid Sequence, pubmed-meshheading:15481977-Animals, pubmed-meshheading:15481977-COS Cells, pubmed-meshheading:15481977-Cattle, pubmed-meshheading:15481977-Humans, pubmed-meshheading:15481977-Hydrogen Bonding, pubmed-meshheading:15481977-Membrane Proteins, pubmed-meshheading:15481977-Models, Molecular, pubmed-meshheading:15481977-Molecular Sequence Data, pubmed-meshheading:15481977-Mutagenesis, Site-Directed, pubmed-meshheading:15481977-Protein Structure, Secondary, pubmed-meshheading:15481977-Purinergic P2 Receptor Agonists, pubmed-meshheading:15481977-Purinergic P2 Receptor Antagonists, pubmed-meshheading:15481977-Radioligand Assay, pubmed-meshheading:15481977-Receptors, Purinergic P2, pubmed-meshheading:15481977-Receptors, Purinergic P2Y1, pubmed-meshheading:15481977-Receptors, Purinergic P2Y12, pubmed-meshheading:15481977-Rhodopsin, pubmed-meshheading:15481977-Sequence Alignment
pubmed:year
2004
pubmed:articleTitle
Architecture of P2Y nucleotide receptors: structural comparison based on sequence analysis, mutagenesis, and homology modeling.
pubmed:affiliation
Molecular Recognition Section, Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892-0810, USA.
pubmed:publicationType
Journal Article