Williams L T, Snyderman R, Pike M C, Lefkowitz R J
Proc Natl Acad Sci U S A. 1977 Mar;74(3):1204-8. doi: 10.1073/pnas.74.3.1204.
Synthetic N-formylmethionyl peptides are chemotactic attractants for human polymorphonuclear leukocytes. The well-defined structure-activity relationship of these peptides in eliciting a chemotactic response suggests that the interaction of the peptides with a specific cellular binding site may initiate chemotaxis. By using tritiated N-formylmethionyl-leucyl-phenylalanine (fMet-Leu-[3H]Phe), a potent chemotactic peptide with high specific radioactivity, we have directly identified binding sites on human polymorphonuclear leukocytes. Binding of fMet-Leu-[3H]Phe to polymorphonuclear leukocytes is rapid (t1/2 less than 2 min) and reversible. The equilibrium dissociation constant (KD) for the interaction of fMet-Leu-[3H-A1Phe with the binding site is 12-14 nM at 37 degrees. The number of binding sites is approximately 2000 per cell. The specificity of the binding sites for a series of N-formylmethionyl peptides exactly reflects the specificity of the chemotactic response to the peptides in that they compete for the binding sites and initiate chemotaxis with the same order of potency (fMet-Leu-Phe greater than fMet-Met-Met greater than fMet-Phe greater than fMet-Leu greater than fMet),fPhe-Met is a competitive antagonist of the chemotactic activity of N-formylmethionyl peptides and has a calculated KD of 6x10-5 M. FPhe-Met also half-maximally inhibits binding of fMet-Leu[3H]Phe binding was the highest in polymorphonuclear leukocytes. No binding of fMet-Leu-[3H]Phe to human erythrocytes could be detected. These data indicate that fMet-Leu-[3H]Phe can be used to identify binding sites for chemotactic peptides on human polymorphonuclear leukocytes. It is likely that these binding sites initiate the specific response of motile cells to N-formylmethionyl peptides.
合成的N-甲酰甲硫氨酰肽是人类多形核白细胞的趋化吸引剂。这些肽在引发趋化反应方面明确的构效关系表明，肽与特定细胞结合位点的相互作用可能启动趋化作用。通过使用具有高比放射性的强趋化肽——氚标记的N-甲酰甲硫氨酰-亮氨酰-苯丙氨酸（fMet-Leu-[³H]Phe），我们直接鉴定了人类多形核白细胞上的结合位点。fMet-Leu-[³H]Phe与多形核白细胞的结合迅速（半衰期小于2分钟）且可逆。在37℃时，fMet-Leu-[³H]Phe与结合位点相互作用的平衡解离常数（KD）为12 - 14 nM。每个细胞的结合位点数约为2000个。一系列N-甲酰甲硫氨酰肽结合位点的特异性准确反映了对这些肽趋化反应的特异性，因为它们竞争结合位点并以相同的效力顺序引发趋化作用（fMet-Leu-Phe＞fMet-Met-Met＞fMet-Phe＞fMet-Leu＞fMet），fPhe-Met是N-甲酰甲硫氨酰肽趋化活性的竞争性拮抗剂，计算得出的KD为6×10⁻⁵ M。FPhe-Met也能半最大程度地抑制fMet-Leu[³H]Phe的结合，在多形核白细胞中的结合最高。未检测到fMet-Leu-[³H]Phe与人红细胞的结合。这些数据表明，fMet-Leu-[³H]Phe可用于鉴定人类多形核白细胞上趋化肽的结合位点。这些结合位点很可能启动运动细胞对N-甲酰甲硫氨酰肽的特异性反应。
