Schreiber R E, Prossnitz E R, Ye R D, Cochrane C G, Jesaitis A J, Bokoch G M
Department of Cell Biology, Scripps Research Institute, La Jolla, California 92037.
J Leukoc Biol. 1993 Apr;53(4):470-4. doi: 10.1002/jlb.53.4.470.
A recombinant human neutrophil N-formyl peptide receptor (rFPR) expressed in transfected mouse fibroblasts (TX2 cells) was analyzed for its ability to couple physically with the heterotrimeric G protein, Gi. Immunoprecipitation of photoaffinity-labeled rFPR and endogenous neutrophil formyl peptide receptor (nFPR) with an anti-FPR peptide antibody demonstrated that the receptors were identical in both size and extent of glycosylation. Coupling of rFPR with endogenous TX2 Gi was demonstrated by coimmunoprecipitation of the two proteins with an anti-Gi antibody. Moreover, rFPR was able to form a physical complex with purified Gi in a soluble reconstitution system. We observed similar affinities of rFPR and nFPR for Gi. This report provides the first direct evidence that rFPR associates physically with Gi and provides a foundation for analysis of the G protein coupling capacity of mutant rFPRs.
对转染小鼠成纤维细胞（TX2细胞）中表达的重组人中性粒细胞N-甲酰肽受体（rFPR）与异源三聚体G蛋白Gi进行物理偶联的能力进行了分析。用抗FPR肽抗体对光亲和标记的rFPR和内源性中性粒细胞甲酰肽受体（nFPR）进行免疫沉淀，结果表明，这两种受体在大小和糖基化程度上是相同的。用抗Gi抗体对这两种蛋白进行共免疫沉淀，证明rFPR与内源性TX2 Gi偶联。此外，rFPR能够在可溶性重组系统中与纯化的Gi形成物理复合物。我们观察到rFPR和nFPR对Gi具有相似的亲和力。本报告首次提供了rFPR与Gi物理缔合的直接证据，并为分析突变型rFPR的G蛋白偶联能力奠定了基础。
