CAJ | 학술논문

目的研究以cGMP为特异底物的磷酸二酯酶5（PDE5）的水解酶活性及PDE5选择性抑制剂西地那非(sildenafil)的抑制作用。方法以牛阴茎海绵体为原料，通过快速蛋白液相系统分离纯化得到PDE同工酶，然后以3H-cGMP为底物，经纯化的PDE同工酶催化水解为3H-GMP，进一步在含5′-核苷酸酶的蛇毒液作用下，脱去5′-磷酸生成3H-鸟苷，在闪烁剂激发下测定3H-鸟苷的变化，绘制PDE活性曲线。以不同剂量的PDE5选择性抑制剂西地那非作用于PDE5，经Dixon Plots法处理结果以观察西地那非的抑制效应。结果由牛阴茎海绵体纯化得到三个PDE同工酶峰，其中第三个峰对cGMP水解活性最强，且西地那非对第三峰抑制作用明显。结论海绵体中的PDE以cGMP-特异的PDE5为主，西地那非是PDE5的选择性抑制剂，通过以上实验观察可初步确定第三峰为PDE5，其结果与文献报道相符。
목적연구이cGMP위특이저물적린산이지매5（PDE5）적수해매활성급PDE5선택성억제제서지나비(sildenafil)적억제작용。방법이우음경해면체위원료，통과쾌속단백액상계통분리순화득도PDE동공매，연후이3H-cGMP위저물，경순화적PDE동공매최화수해위3H-GMP，진일보재함5′-핵감산매적사독액작용하，탈거5′-린산생성3H-조감，재섬삭제격발하측정3H-조감적변화，회제PDE활성곡선。이불동제량적PDE5선택성억제제서지나비작용우PDE5，경Dixon Plots법처리결과이관찰서지나비적억제효응。결과유우음경해면체순화득도삼개PDE동공매봉，기중제삼개봉대cGMP수해활성최강，차서지나비대제삼봉억제작용명현。결론해면체중적PDE이cGMP-특이적PDE5위주，서지나비시PDE5적선택성억제제，통과이상실험관찰가초보학정제삼봉위PDE5，기결과여문헌보도상부。
Purpose　The aim is to study the activity and the sildenafil selective inhibition of PDE5. Methods　The PDE isoenzymes were purified from bovine penis corpus cavernosum tissue by FPLC system. PDE activity was assayed by using 3　H-cGMP as substrate, the PDE isoenzymes hydrolyzed it to 3　H-GMP, and 3　H-GMP was further hydrolyzed to 3　H-guanosine by 5′-nuclease of snake venom. Add scintillation cocktail to observe the PDE isoenzymes activity. The selective inhibitor sildenafil of different concentrations were used to observe the inhibition of PDE5. Data replotted according to procedure of Dixon plots.Results　Three PDE isoenzyme peaks were purificated from bovine corpus cavernosum. The PDE of the third peak had the strongest activity of cGMP hydrolyzation which could be inhibited by sildenafil apparently.Conclusion　Since PDE5 was mainly found in corpus cavernosum tissue of mammalian, and sildenafil was a selective inhibitor of PDE5. It was suggested that the third peak was PDE5. The result was in agreement with the article reported.