Characterization of the binding site on the formyl peptide receptor using three receptor mutants and analogs of Met-Leu-Phe and Met-Met-Trp-Leu-Leu
Characterization of the binding site on the formyl peptide receptor using three receptor mutants and analogs of Met-Leu-Phe and Met-Met-Trp-Leu-Leu
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DOI：
10.1074/jbc.m003081200
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发表时间：
2000-12-15
影响因子：
4.8
通讯作者：
Jesaitis, AJ
中科院分区：
文献类型：
--
作者：
Mills, JS;Miettinen, HM;Jesaitis, AJ
甲酰肽受体（FPR）是一种在吞噬细胞表面发现的趋化性G蛋白偶联受体。我们先前已经表明，甲酰基肽结合位点映射到跨膜区（Miettinen，H. R.，米尔斯，J.S.，Gripentrog，J.M.，Dratz，E.一、格兰杰，E. L.，和Jesaitis，A. J.（1997）J. Immunol.，159，4045 - 4054）。最近的报道已经表明，非甲酰化的肽，例如MMWLL也可以激活这种受体（Chen，J.，伯恩斯坦，H.美国，陈美，Wang，L.，美国，Ishi，M.，图尔克角W.，和Coughlin，S. R.（1995）J，Biol·Chem.270，23398 - 23401）。在这里，我们表明，不同的NH2-末端类似物的MMWLL或MLF的结合的选择性可以显着改变突变天冬酰胺或精氨酸-201丙氨酸的天冬氨酸-106。D106 N和R201 A在配体特异性方面产生了类似的变化，包括结合HIV-I肽DP178的能力增强。相比之下，突变R205A在fMLF的COOH末端表现出改变的特异性，其中R205A结合fMLF-O-丁基> fMLF-O-甲基> fMLF，而wt FPR结合fMLF> fMLF-O-甲基，类似于fMLF-O-丁基。这些数据与我们先前发现的fMLF的亮氨酸侧链可能在FPR（VRK 95）-V-93附近与FPR结合（米尔斯，J.S.，Miettinen，H. M.，Barnidge，D.，Vlases，M. J.，Wimer-Mackin，S.，Dratz，E.一、和Jesaitis，A. J.（1998）J.Biol.Chem.273，10428 - 10435），表明fMLF在EPR结合口袋中最可能的定位大致平行于第五个跨膜螺旋，其中fMLF的甲酰胺基团与Asp-106和Arg-201氢键结合，亮氨酸侧链指向第二个跨膜区，并且fMLF的COOH末端羧基与Arg-205离子配对。
The formyl peptide receptor (FPR) is a chemotactic G protein-coupled receptor found on the surface of phagocytes. We have previously shown that the formyl peptide binding site maps to the membrane-spanning region (Miettinen, H. R., Mills, J. S., Gripentrog, J. M., Dratz, E. A., Granger, E. L., and Jesaitis, A. J. (1997) J. Immunol, 159, 4045-4054). Recent reports have indicated that nonformylated peptides, such as MMWLL can also activate this receptor (Chen, J., Bernstein, H. S., Chen, M.,Wang, L., Ishi, M., Turck, C. W., and Coughlin, S. R. (1995) J, Biol. Chem. 270, 23398-23401.) Here we show that the selectivity for the binding of different NH2-terminal analogs of MMWLL or MLF can be markedly altered by mutating Asp-106 to asparagine or Arg-201 to alanine. Both D106N and R201A produced a similar change in ligand specificity, including an enhanced ability to bind the HIV-I peptide DP178. In contrast, the mutation R205A exhibited altered specificity at the COOH terminus of fMLF, with R205A binding fMLF-O-butyl > fMLF-O-methyl > fiMLF, whereas wt FPR bound fMLF > fMLF-O-methyl similar to fMLF-O-butyl. These data, taken together with our previous finding that the leucine side chain of fMLF is probably bound to FPR near FPR (VRK95)-V-93 (Mills, J. S., Miettinen, H. M., Barnidge, D., Vlases, M. J., Wimer-Mackin, S., Dratz, E. A., and Jesaitis, A. J. (1998) J. Biol. Chem. 273, 10428-10435.), indicate that the most Likely positioning of fMLF in the binding pocket of EPR is approximately parallel to the fifth transmembrane helix with the formamide group of fMLF hydrogen-bonded to both Asp-106 and Arg-201, the leucine side chain pointing toward the second transmembrane region, and the COOH-terminal carboxyl group of fMLF ion-paired with Arg-205.