聚乙二醇化重组人生长激素的性质及活性

目的　研究聚乙二醇化重组人生长激素的性质及活性。方法　通过分子筛高效液相 (SizeexclusionHPLC)、毛细管等电聚焦 (CIEF)、圆二色谱 (CD)以及荧光光谱等方法检测单聚PEG- GH的纯度、等电点、二级结构以及分子构象 ;采用妊娠兔肝细胞放射受体分析法 (RRA)检测单聚PEG GH的体外受体亲和力 ,采用未成年去垂体大鼠体重增加法检测体内生物活性。结果　分子筛高效液相图谱上仅出现一个峰 ,表明样品纯度很高 ,等电点范围为 5 .1～ 5 .2 ,与未修饰的rhGH的等电点相一致 ;CD谱与荧光光谱均表明单个mPEG ALD的修饰对rhGH的二级结构以及分子构象没有明显影响 ;与未修饰的rhGH相比 ,单聚PEG GH体外受体亲和力明显降低 ;但体内作用时间更长 ,且更加有效。结论　此结果为进一步研究和开发长效重组人生长激素提供了理论依据。

Characteristics and Biological Activity of Mono-PEG-GH

Objective To study the charactristics and bio logi cal activity of PEG-modified recombinant human growth hormone(mono-PEG-GH). Methods The purity,isoelectric point,secondary structure and mole cular conformation of mono-PEG-GH were analyzed by the methods including size exclusion HPLC,capillary isoelectric focusing,circular dichrosim(CD) and fluores cence spectrometry.The receptor binding activity in vitro of mono-PEG-GH w as e valuated by analyzing its inhibitory effect on the binding of 125I-hG H to growt h hormone receptor on the membrane of pregnant rabbit liver cells using radiorec eptor assay.The biological activity in vivo of mono-PEG-GH was analyzed by bod y weight gain test in hypophysectomized rats.Results High purit y of mono-PEG-GH was proved by the only peak on size exclusion HPLC profile.The isoelectric point of mono-PEG-GH was 5 1-5 2,which was consistent with tha t of unmodified rhGH.Neither CD nor fluorescence spectrometry showed the influen ce of modification with mPEG-ALD on the secondary structure or molecular confor mation of rhGH.Compared with that of unmodified rhGH,the receptor affinity in vi tro of mono-PEG-GH decreased significantly.However,its receptor affinity i n vivo was more persistent and effective.Conclusion It provid ed a theoretical basis for the study and development of long-acting rhGH. [