Identification of residues responsible for ligand recognition and regioisomeric selectivity of lysophosphatidic acid receptors expressed in mammalian cells

Yuko Fujiwara, Vineet Sardar, Akira Tokumura, Daniel Baker, Kimiko Murakami-Murofushi, Abby Parrill, Gabor Tigyi

Research output: Contribution to journalArticle

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Abstract

The endothelial differentiation gene family encodes three highly homologous G protein-coupled receptors for lysophosphatidic acid (LPA). Based on baculoviral overexpression studies, differences have been proposed in the structure-activity relationship (SAR) of these receptors. We have compared the SAR of the individual receptors either overexpressed transiently at high or at lower levels following stable transfection in LPA-nonresponsive RH7777 cells. The SAR in transfected RH7777 cells was markedly different from that described in insect cells. The LPA 3 receptor has been pro posed to be selectiveiy activated by unsaturated LPA species and shows a strong preference for sn-2 versus the sn-1 acyl-LPA regioisomer. Because of the short half-life of sn-2 LPA due to acyl migration under some conditions, we have synthesized acyl migration-resistant analogs using an acetyl group in place of the free hydroxyl group in order to evaluate LPA receptor SAR. Only LPA 1 and LPA 2 showed regioisomeric preference and only for the 18:2 fatty acyl-stabilized LPA Sn-1 regioisomer. To identify residues involved in ligand recognition of LPA 3 , we developed and validated computational models of LPA 3 complexes with the analogs studied. The models revealed that Arg-3.28 and Gln-3.29 conserved within the LPA-selective endothelial differentiation gene receptors and the more variable Lys-7.35 and Arg-5.38 Of LPA 3 form critical interactions with the polar head-group of LPA. The models identified Leu-2.60 and Val-7.39of LPA 3 underlying the regioisomer-selective interaction with the acetyl group of the stabilized regioisomers. Mutation of Leu-2.60 to alanine selectively increased the EC 50 Of the sn-2 acetyl-LPA regioisomers, whereas alanine replacement of Val-7.39 profoundly affected both regioisomers.

Original languageEnglish (US)
Pages (from-to)35038-35050
Number of pages13
JournalJournal of Biological Chemistry
Volume280
Issue number41
DOIs
StatePublished - Oct 14 2005

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Lysophosphatidic Acid Receptors
Cells
Ligands
Structure-Activity Relationship
lysophosphatidic acid
Alanine
Genes

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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Identification of residues responsible for ligand recognition and regioisomeric selectivity of lysophosphatidic acid receptors expressed in mammalian cells. / Fujiwara, Yuko; Sardar, Vineet; Tokumura, Akira; Baker, Daniel; Murakami-Murofushi, Kimiko; Parrill, Abby; Tigyi, Gabor.

In: Journal of Biological Chemistry, Vol. 280, No. 41, 14.10.2005, p. 35038-35050.

Research output: Contribution to journalArticle

Fujiwara, Yuko ; Sardar, Vineet ; Tokumura, Akira ; Baker, Daniel ; Murakami-Murofushi, Kimiko ; Parrill, Abby ; Tigyi, Gabor. / Identification of residues responsible for ligand recognition and regioisomeric selectivity of lysophosphatidic acid receptors expressed in mammalian cells. In: Journal of Biological Chemistry. 2005 ; Vol. 280, No. 41. pp. 35038-35050.
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AU - Fujiwara, Yuko

AU - Sardar, Vineet

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AU - Baker, Daniel

AU - Murakami-Murofushi, Kimiko

AU - Parrill, Abby

AU - Tigyi, Gabor

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