Direct regulation of GTP-binding regulatory proteins by biologically active peptides.

研究了从黄蜂毒液中分离的两亲肽P物质（SP）和从哺乳动物中分离的神经肽P物质（MP）诱导大鼠腹膜肥大细胞胞吐的机制，以阐明MP和SP是否通过直接激活这些细胞中gtp结合调节蛋白（G蛋白）而引起胞吐。MP和SP在1-30 muM浓度下诱导肥大细胞释放非溶解性β -己糖氨酸酶，百日咳毒素阻止了这些作用，表明百日咳毒素敏感的G蛋白参与了分泌。细胞外Ca^<2+>的存在对于肽诱导的分泌并不是必需的，但它增加了分泌的动力学和最大反应，并且其存在降低了效力。用Ala取代MP的疏水氨基酸残基显著降低了β -己糖氨酸酶释放的刺激和G_i的激活。MP需要赖氨酸残基来刺激β -己糖氨酸酶释放并激活G_i。这些结果表明疏水和带正电的侧链对于MP在体外刺激肥大细胞和G_i中的靶分子的重要性。疏水和带正电的氨基酸残基也需要SP激活肥大细胞和G_i。[Lys^<10>, Leu^<13>] MP是刺激β -己糖氨酸酶释放而不引起细胞裂解的最有效的类似物，表明该肽是肥大细胞最有用的化学刺激剂。然而，该肽仅轻微激活G_i。基于腹膜肥大细胞胞外分泌与MP、SP及其类似物体外激活G_i蛋白的构效关系，讨论了MP和SP直接激活腹膜肥大细胞中G_i样蛋白的可能性。

The mechanisms of exocytosis from rat peritoneal mast cells induced by mastoparan (MP) and substance P (SP), an amphiphilic peptide isolated from wasp venom, and a neuropeptide isolated from mammals, respectively, were investigated to elucidate whether MP and SP cause exocytosis to directly activate GTP-binding regulatory proteins (G proteins) in these cells. MP and SP induced non-lytic beta-hexosaminidase release from mast cells at concentrations of 1-30 muM,and these effects were prevented by pertussis toxin, indicating the involvement of pertussis toxin-sensitive G proteins in the secretion. The presence of extracellular Ca^<2+> was not essential for peptide-induced secretion, but it increased the kinetics and maximal response of the secretion, and the potency was decreased in its presence. Replacing the hydrophobic amino acid residues of MP with Ala significantly decreased the stimulation of beta-hexosaminidase release and the activation of G_i. Lys residue (s) was required for MP to stimulate beta-hexosaminidase release and activate G_i. These results demonstrated the importance of hydrophobic and positively charged side chains for MP to stimulate target molecules in mast cells and G_i in vitro. Both hydrophobic and positively charged amino acid residues were also required the activation of mast cells and G_i by SP.[Lys^<10>, Leu^<13>] MP was the most potent analog that stimulated beta-hexosaminidase release without causing cell lysis, indicating that this peptide is the most useful chemical stimulator of the mast cells. However, this peptide only slightly activated G_i. The possibility of direct activation of a G_i like protein by MP and SP in peritoneal mast cells in discussed based upon the structure-activity correlation between exocytosis from these cells and G_i-protein activation in vitro by MP,SP and their analogs.