Linked Life Data

15035619

Source:http://linkedlifedata.com/resource/pubmed/id/15035619

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
12
pubmed:dateCreated
2004-3-23
pubmed:abstractText
Oxytocin receptor (OTR) activates the GTP-binding protein Galpha(q). To investigate whether the N-terminal region of the fourth intracellular domain of this receptor, which forms putative helix 8, plays a role in coupling, its hydrophilic residues (H7.59, H7.62, E7.63, Q7.66, and R7.67) were mutated individually to alanine. In COSM6 cells, these mutants were expressed at equivalent concentrations, but at lower concentrations than OTR. Alanine substitution for H7.62 or Q7.66 did not substantially affect the affinity for OT (K(d) = 0.63 and 0.48 nM, respectively, vs 0.52 nM for the wild type), whereas substitution for H7.59, E7.63, or R7.67 reduced the affinity 5-6-fold. When expressed at equal concentrations, OTR-H7.62/A and OTR-Q7.66/A stimulated phosphatidylinositide turnover as well as OTR, whereas OTR-H7.59/A, OTR-E7.63/A, and OTR-R7.67/A exhibited an impaired ability to respond to OT. Therefore, residues H7.59, E7.63, and R7.67 within the putative hydrophilic interface appeared to influence both the OTR conformation and Galpha(q) coupling. To explore this further, five multiple alanine substitution mutants were constructed. Alanine modification at H7.62 and Q7.66 did not substantially affect the affinity for OT (K(d) = 0.75 nM), whereas any combination of alanine substitutions for H7.59, E7.63, and R7.67 produced mutant receptors that lost high-affinity ligand binding. While OTR-(H7.62,Q7.66)/A exhibited PLC activation equivalent to that of OTR, receptors with two or more changes in H7.59, E7.63, and R7.67 lost the ability to respond to OT in a dose-dependent manner. Five residues (L7.60, F7.61, L7.64, V7.65, and F7.68) in the opposite hydrophobic interface were also mutated to alanine. None of these substitutions affected ligand binding; only OTR-(L7.60,F7.61)/A had a somewhat weaker ability to activate PLC. These data are consistent with the prediction that these residues lie within an amphipathic alpha-helix and emphasize the importance of this hydrophilic interface, and particularly of H7.59, E7.63, and R7.67, in OTR function.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Mar
pubmed:issn
0006-2960
pubmed:author
pubmed:issnType
Print
pubmed:day
30
pubmed:volume
43
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
3490-8
pubmed:dateRevised
2010-11-18
pubmed:meshHeading
pubmed-meshheading:15035619-Alanine, pubmed-meshheading:15035619-Amino Acid Sequence, pubmed-meshheading:15035619-Amino Acid Substitution, pubmed-meshheading:15035619-Animals, pubmed-meshheading:15035619-Arginine, pubmed-meshheading:15035619-COS Cells, pubmed-meshheading:15035619-Cattle, pubmed-meshheading:15035619-Glutamic Acid, pubmed-meshheading:15035619-Histidine, pubmed-meshheading:15035619-Humans, pubmed-meshheading:15035619-Hydrophobic and Hydrophilic Interactions, pubmed-meshheading:15035619-Molecular Sequence Data, pubmed-meshheading:15035619-Oxytocin, pubmed-meshheading:15035619-Protein Binding, pubmed-meshheading:15035619-Protein Structure, Secondary, pubmed-meshheading:15035619-Rats, pubmed-meshheading:15035619-Receptors, Oxytocin, pubmed-meshheading:15035619-Receptors, Vasopressin, pubmed-meshheading:15035619-Signal Transduction, pubmed-meshheading:15035619-Transfection
pubmed:year
2004
pubmed:articleTitle
Residues in the hydrophilic face of putative helix 8 of oxytocin receptor are important for receptor function.
pubmed:affiliation
Department of Biomedical Sciences, Colorado State University, Fort Collins, Colorado 80523, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, Non-U.S. Gov't