人碳酸酐酶Ⅱ(hCAⅡ)抑制剂筛选技术及应用  

Screening of new human carbonic anhydrase Ⅱ(hCA Ⅱ) inhibitors

作　　者：肖忠海[1] 段瑞峰[1] 崔文玉[2] 汪海[1,2,3] 

机构地区：[1]军事医学科学院卫生学环境医学研究所,天津300050 [2]军事医学科学院毒物药物研究所,北京100850 [3]北京赛德维康医药研究院,北京100039

出　　处：《中国药理学通报》2010年第2期182-185,共4页Chinese Pharmacological Bulletin

基　　金：国家重大新药创制科技重大专项(No2009ZXJ09004-086)

摘　　要：目的建立人碳酸酐酶Ⅱ(hCAⅡ)抑制剂筛选技术,并用于评价新hCAⅡ抑制剂。方法以大肠杆菌表达的hCAⅡ作为酶源,按照酶反应体系构建原则,在pH7.6和25℃的条件下,以碳酸酐酶分解对硝基酚乙酸酯引起348nm处光吸收增加来计算酶活力。在确定碳酸酐酶抑制剂筛选条件后,对35个靶向碳酸酐酶的新化合物进行了hCAⅡ的抑酶活性评价。结果采用大肠杆菌表达的hCAⅡ为酶源,成功建立了hCAⅡ抑制剂筛选技术,该技术具有可靠、快速、简便的优点,发现10个新化合物对hCAⅡ的抑制率大于醋氮酰胺,9个新化合物对hCAⅡ的抑制率与醋氮酰胺相当。结论hCAⅡ抑制剂筛选技术可用于hCAⅡ抑制剂的大规模筛选,为发现高效、高亚型选择性的碳酸酐酶靶向性药物奠定基础。进一步深入研究这19个化合物有望获得高效、低副作用的急性高原病防治药物。Ahn To provide practical method for screening human carbonic anhydrase Ⅱ （hCA Ⅱ ） inhibitors in drug discovery. Methods hCA Ⅱ protein was obtained from induced BL21 （DE3） E. coli containing plasmid pET-28b-hCA Ⅱ. The hCA Ⅱ activity was detected under pH 7.6 and 25℃; by its esterase activity which could decompose PNPA to increase the photoabsorption at 348 nm. After the assay conditions were finally selected, 35 new compounds were tested. Results A practical method for screening hCA Ⅱ inhibitors was successfully constituted by using recombinant hCA Ⅱ protein expressed in E. coli as the sourceof hCA H enzyme. 10 new compounds had better inhibitory effect and 9 new compounds had the same inhibitory effect on hCA Ⅱ compared with acetazolamide. Conclusions The hCA Ⅱ inhibitor screening technique constituted in this work possesses advantages of being reliable, rapid, and practical. 19 new compounds are worth further research for developing high efficiency and low side effect drugs used for high-altitude illness.

关 键 词：急性高原病 醋氮酰胺 抑制剂 碳酸酐酶 同工酶 原核表达 筛选技术 

分 类 号：R345.72[医药卫生—基础医学] R594.305