知识与财富的链接
目的:为了寻找高活性和长半衰期生的GHRH类似肽。方法:通过使用独特的酸敏感水解位点Asp-Pro的原核表达系统,构建了新的Pro-hGHRH(1-44)-Gly-Gly-Cys类似肽。通过重组细菌裂解、包含体洗涤、乙醇分级沉淀、酸水解、SP-Sephadex C-25和Sephadex G-10柱层析等技术,纯化了高纯度的Pro-hGHRH(1-44)-Gly-Gly-Cys肽。通过使用SDS-PAGE、离子化质谱、雌大鼠垂体和人流产胎儿垂体,测定了多肽的纯度、分子量、生长激素释放活性。结果:Pro-hGHRH(1-44)-Gly-Gly-Cys肽分子量5373Da与实际值吻合,0.1~10 μg/ml的肽剂量不论是对人垂体还是大鼠垂体都增加了垂体生长激素的释放,大鼠垂体生长激素的释放具有剂量依赖性。与标准的hGHRH(1-40)肽比较,新的类似肽有较高的GH释放活性。结果也显示了,Pro-GHRH(1-44)-Gly-Gly-Cys与Pro-hGHRH(1-44)肽的GH释放活性无统计学差异。结论：新的类似肽有较好的生长激素释放活性、功能选择性和种属特异性。
Objective:Growth hormone releasing hormone(GHRH)is one of the hypothalamus hormones.Because of its potential applications in agriculture and human medicine,an analog with high GHRH activity and prolonged half-life has been looked for.Methods:By using the fusion protein expression protocol with unique acid labile linker Asp-Pro,the Pro-hGHRH(1-44)-Gly-Gly-Cys peptide was obtained.The peptide was purified to homogeneity by means of cell disruption,washing of inclusion body,ethanol fractionation precipitation,acid hydrolysis,SP-Sephadex C-25 and Sephadex G-10 column chromatography.Results:The peptide molecular weight of 5373 Da measured by EIS-MS is coincident with the actual values and its purity was determined by SDS-PAGE.The peptide at 0.1～10μg/ml increased GH releases from both human pituitary and rat pituitary.The rat pituitary GH-releasing activity increased in a dose-dependent manner.Compared with the standard hGHRH(1-40),the GHRH analog had higher rGH-releasing activity.The standard hGHRH(1-40)at 0.01～2.0μg/ml showed no measured rat pituitary GH release.The activity comparisons indicated that the rat pituitary GH-releasing activity of the Pro-GHRH(1-44)Gly-Gly-Cys peptide was not more than that of the Pro-hGHRH(1-44)peptide,whereas the human pituitary GH-releasing activity of the Pro-GHRH(1-44)-Gly-Gly-Cys peptide was far less than that of the Pro-hGHRH(1-44)peptide despite there was no significance between them.Conclusion:The activity results indicated that the novel GHRH analog had good GH-releasing activity,function selectivity and species specificity.
