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利用黑棘鲷 (Acanthopagrusbutcheri)生长激素 (brGH )作为配体 ,建立了黑鲷 (Sparusmacrocephalus)生长激素受体放射受体分析法 (RadioreceptorAssay ,RRA)并分析其组织分布特征。在 2 5℃下 ,12 5I brGH与黑鲷肝细胞膜蛋白的特异性结合具有如下特点 :(1)时间依赖性 特异性结合达最大结合量一半的时间 (Ta1/ 2 )为 10 4h ,结合达到平衡状态需 2 0h ;激素 受体复合物的解离在 3h之内最快 ,2 4h后仅解离约40 % ,表明激素 -受体复合物的结合仅具部分可逆转性 ;(2 )可饱和性 12 5I brGH与黑鲷肝细胞膜蛋白的特异性结合随着膜蛋白浓度或12 5I brGH加入量的上升而呈逐渐上升并达到饱和状态的趋势 ;(3)可取代性 12 5I brGH与黑鲷肝细胞膜蛋白的特异性结合可被非标记brGH竞争性地取代 ,将特异性结合取代 5 0 % (ED50 )所需非标记brGH约为 3 8ng ,非标记重组金鱼生长激素 (rgfGH)及人生长激素 (hGH)对12 5IbrGH的竞争性取代能力明显较brGH低 ,而重组金鱼催乳素 (rgfPRL)、羊催乳素 (oPRL)在该系统中几乎无交叉反应。Scachard作图分析表明黑鲷肝细胞膜蛋白存在brGH单一的高亲和结合位点 ,Ka为 (3 49± 0 2 4)× 10 10 (mol/L) -1,Bmax为 146 8± 10 6fmol/mgprotein。结果表明 ,brGH与黑鲷肝细胞膜蛋白的?
Using Acanthopagrus pubescens growth hormone (brGH) as ligand, the Radioreceptor Assay (RRA) of Sparus macrocephalae growth hormone receptor was established and its tissue distribution characteristics were analyzed. The specific binding of 125I brGH to the hepatocyte membrane protein of black sea bream at 25 ° C has the following characteristics: (1) the time-dependent specific binding (Ta1 / 2) for half of the maximum bound amount is 10 4 h, The equilibrium state required 20h; the dissociation of hormone receptor complex was the fastest within 3h and only 40% dissociated after 24 hours, indicating that the binding of hormone-receptor complex was only partially reversible; (2) Saturability 12 5I brGH and black sea bream hepatocyte membrane protein specific binding with the membrane protein concentration or 12 5I brGH the amount of the increase was gradually increased and reached a saturation trend; (3) substitutable 12 5I brGH and The specific binding of hepatocyte membrane proteins in black sea bream can be competitively replaced by non-labeled brGH, which will require about 38 ng of non-labeled brGH to specifically bind to the ED50, and 35 mg of unlabeled recombinant goldfish growth hormone (rgfGH) and Competitive replacement of human growth hormone (hGH) with 125IbrGH was significantly lower than that of brGH, whereas recombinant goldfish prolactin (rgfPRL) and sheep prolactin (oPRL) had almost no cross-reaction in this system. Scachard mapping analysis showed that there was a single high affinity binding site of brGH in membrane proteins of black sea bream, Ka was (3 49 ± 0 2 4) × 10 10 (mol / L) -1 and Bmax was 146 8 ± 10 6 fmol / mgprotein. The results showed that brGH and black sea bream liver cell membrane protein?