Mackin W M, Huang C K, Becker E L
J Immunol. 1982 Oct;129(4):1608-11.
F Met-Leu-[3H]Phe and f Nle-Leu-[3H]Phe binding to rabbit peritoneal neutrophils and purified membranes were measured at 4 degrees C silicone oil centrifugation assays, and the results were analyzed by the LIGAND computer program, which permits analysis of ligand binding to multiple classes of binding sites. LIGAND analysis of peptide binding to intact neutrophil indicated that both f Met-Leu-[3H]Phe and f Nle-Leu-[3H]Phe detected two population of binding sites. The apparent Kd values for f Met-Leu-[3H]Phe binding were 1.6 +/- 1.0 X 10(-9) M and 2.2 +/- 0.9 X 10(-8) M, respectively, and 3.1 +/- 0.2 X 10(-9) M and 1.2 +/- 0.6 X 10(-7) M for f Nle-Leu-[3H]Phe. Furthermore, the higher affinity sites detected on whole cells comprised approximately 15 to 30% of the total sites. Two populations of binding sites were also detected on purified neutrophil plasma membranes by both radiolabeled chemotactic peptides. LIGAND analysis of peptide binding to purified membranes yielded apparent Kd values of 5.0 +/- 2.5 X 10(-10) M and 4.8 +/- 0.6 X 10(-8) M for f Met-Leu-[3H]Phe binding, and 4.7 +/- 4.2 X 10(-10) M and 3.0 +/- 1.3 X 10(-8) M for f Nle-Leu-[3H]Phe. The percentage of higher affinity sites detected by f Met-Leu-[3H]Phe and f Nle-Leu-[3H]Phe on purified membranes were 1 to 5% of the total sites detected. These data are consistent either with the existence of two independent binding sites for formylpeptides on rabbit neutrophils or receptor negative cooperativity.
在4℃下，通过硅油离心分析法测定了F Met-Leu-[3H]苯丙氨酸和f Nle-Leu-[3H]苯丙氨酸与兔腹膜中性粒细胞及纯化膜的结合情况，并使用LIGAND计算机程序对结果进行分析，该程序可分析配体与多类结合位点的结合情况。对完整中性粒细胞肽结合的LIGAND分析表明，F Met-Leu-[3H]苯丙氨酸和f Nle-Leu-[3H]苯丙氨酸均检测到两类结合位点。F Met-Leu-[3H]苯丙氨酸结合的表观解离常数（Kd）值分别为1.6±1.0×10⁻⁹ M和2.2±0.9×10⁻⁸ M，f Nle-Leu-[3H]苯丙氨酸的Kd值分别为3.1±0.2×10⁻⁹ M和1.2±0.6×10⁻⁷ M。此外，在全细胞上检测到的高亲和力位点约占总位点的15%至30%。两种放射性标记的趋化肽在纯化的中性粒细胞膜上也检测到两类结合位点。对纯化膜肽结合的LIGAND分析得出，F Met-Leu-[3H]苯丙氨酸结合的表观Kd值为5.0±2.5×10⁻¹⁰ M和4.8±0.6×10⁻⁸ M，f Nle-Leu-[3H]苯丙氨酸的Kd值为4.7±4.2×10⁻¹⁰ M和3.0±1.3×10⁻⁸ M。F Met-Leu-[3H]苯丙氨酸和f Nle-Leu-[3H]苯丙氨酸在纯化膜上检测到的高亲和力位点占总检测位点的1%至5%。这些数据要么与兔中性粒细胞上存在两种独立的甲酰肽结合位点一致，要么与受体负协同性一致。
