Biochemical and Biophysical Research Communications
Identification of membrane-type receptor for bile acids (M-BAR)
Section snippets
Materials and methods
Cloning of human BG37. Putative 5′ and 3′ ends of BG37 ORF were identified by RACE method using human fetus Marathon-Ready cDNA (Clontech). Using primers, 5′-CCCCTGTCCCCAGGACCAAGATG-3′ and 5′-TTAGTTCAAGTCC-AGGTCGACACTGCTTT-3′, determined from the RACE products, hBG37 ORF was amplified. PCR conditions were 94 °C for 9 min, followed by 26 cycles of 94 °C for 30 s, 68 °C for 3 min, and finally 94°C for 30 s, then 62 °C for 8 min. The ORFs of mouse and rat BG37 were similarly identified.
Cell culture.
Results and discussion
In a search of the GenBank DNA database with amino acid sequences of known GPCRs, we found a novel human GPCR, human BG37 (hBG37). The putative coding sequence of hBG37 cloned by PCR is 993 bp and encodes 330 amino acids constituting seven transmembrane domains, which is characteristic of GPCRs (Fig. 1). It showed 28% identity to human EDG-1 (sphingosine-1-phosphate receptor), a member of the class 1 (rhodopsin-like) GPCR superfamily. Mouse and rat BG37 cDNAs were also identified, showing high
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