Functional reconstitution of fMet-Leu-Phe receptor in Xenopus laevis oocytes

W. D. Coats, J. Navarro

Research output: Contribution to journalArticle

9 Scopus citations

Abstract

fMet-Leu-Phe (fMLP) receptors were functionally reconstituted into Xenopus laevis oocytes by microinjection with RNA isolated from promyelocytic leukemia cells (HL-60) differentiated with 750 μM N6, O2-dibutyryl cyclic adenosine 3',5'-monophosphate. fMLP-induced Ca2+ mobilization was monitored by measurement of photon emission elicited by aequorin coinjected with RNA into albino X. laevis oocytes. Maximal expression of the fMLP receptor was achieved 48 h after microinjection of RNA. Dose-response experiments revealed a K0.5 of 9.5 nM fMLP which is in good agreement with the dissociation constants of the fMLP receptor complex in human neutrophils. Furthermore, the fMLP-induced Ca2+ mobilization in Xenopus oocytes was blocked by the fMLP receptor inhibitor t-butoxycarbonyl-Met-Leu-Phe. Size fractionation of the RNA and microinjection of the individual fractions indicated that messenger RNA for the fMLP receptor is between 1.5 and 2.0 kilobases. Reconstitution of the fMLP receptor into Xenopus oocytes can be employed to isolate the cDNA encoding the fMLP receptor as well as to study the regulation of the fMLP receptor in a functional system.

Original languageEnglish (US)
Pages (from-to)5964-5966
Number of pages3
JournalJournal of Biological Chemistry
Volume265
Issue number11
StatePublished - May 16 1990
Externally publishedYes

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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