Development of Neutrophile Regulatory Peptides with Receptor Association Structures

中性粒细胞的趋化、脱颗粒和产生超氧阴离子等功能受多种细胞外激动剂的调节，如N-甲酰-甲硫基-亮氨酰-苯丙氨酸(FMLP)。甲酰肽受体(FPR)和甲酰肽受体样蛋白1(FPRL1)是由7种跨膜蛋白组成的超家族，作为fMLP结合受体表达于人中性粒细胞上。我们发现，经人甲酰肽受体跨膜多肽(HFPRTM)处理的人中性粒细胞在fMLP刺激下可增强超氧阴离子的产生。然而，没有鉴定出与hFPRTM多肽相互作用的膜蛋白。为探讨TM多肽对中性粒细胞促增殖活性的序列依赖性，采用固相法和Fmoc化学方法合成了含有甲酰肽受体(FPR)、FPRL1受体和GABA受体TM序列的多肽。用高效液相色谱和MALDI-TOF-MS对合成多肽的结构和结构进行了鉴定，并以人中性粒细胞超氧化物歧化产物的形式进行了生物活性测定。用中性粒细胞激动剂fMLP处理中性粒细胞后，中性粒细胞产生2-3倍的超氧阴离子。来自GABA受体的TM多肽没有明显的启动活性。这些结果表明，TM肽对中性粒细胞的启动作用具有序列依赖性，需要与FPR相关的序列。

Neutrophil functions including chemotaxis, degranulation, and generation of superoxide anion are modulated by diverse extracellular agonists such as N-formyl-methionyl-leucyl-phenylalanine (fMLP). Formyl peptide receptor (FPR) and formyl peptide receptor-like 1 (FPRL1), a superfamily of seven transmembrane (TM) proteins, are expressed on human neutrophils as the fMLP binding receptors. We found that human neutrophils pretreated with human formyl peptide receptor transmembrane (hFPRTM) peptides were enhanced superoxide anion production when stimulated with fMLP. However, a membrane protein interacting with hFPRTM peptides is not identified. To explore the sequence dependences of the TM peptides for neutrophil proming activities, peptides possess the TM sequences of formyl peptide receptor (FPR), FPR like 1 receptor (FPRL1), and GABA receptor were synthesized by solid-phase method with Fmoc chemistry. Homogeneities and structures of synthetic peptides were confirmed by HPLC and MALDI-TOF MS. The biological activities of synthetic peptides were carried out as superoxide production for human neutrophils. Neutrophils treated with TM peptides from FPR and FPRL1 produced 2-3 folds of superoxide anion by the treatment of fMLP, neutrophil agonist. TM peptide from GABA receptor exhibited no significant priming activity. These results suggested the priming effect of TM peptide for neutrophil was sequence dependence and it required the FPR related sequences.