CAJ | 학술논문

目的建立人碳酸酐酶Ⅱ(hCA Ⅱ)抑制剂筛选技术,并用于评价新hCA Ⅱ抑制剂.方法以大肠杆菌表达的hCA Ⅱ作为酶源,按照酶反应体系构建原则,在pH 7.6和25℃的条件下,以碳酸酐酶分解对硝基酚乙酸酯引起348 nm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hCA Ⅱ的抑酶活性评价.结果采用大肠杆菌表达的hCA Ⅱ为酶源,成功建立了hCA Ⅱ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hCA Ⅱ的抑制率大于醋氮酰胺,9个新化合物对hCA Ⅱ的抑制率与醋氮酰胺相当.结论 hCA Ⅱ抑制剂筛选技术可用于hCA Ⅱ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.
목적 건립인탄산항매Ⅱ(hCA Ⅱ)억제제사선기술,병용우평개신hCA Ⅱ억제제.방법 이대장간균표체적hCA Ⅱ작위매원,안조매반응체계구건원칙,재pH 7.6화25℃적조건하,이탄산항매분해대초기분을산지인기348 nm처광흡수증가래계산매활력.재학정탄산항매억제제사선조건후,대35개파향탄산항매적신화합물진행료hCA Ⅱ적억매활성평개.결과 채용대장간균표체적hCA Ⅱ위매원,성공건립료hCA Ⅱ억제제사선기술,해기술구유가고、쾌속、간편적우점,발현10개신화합물대hCA Ⅱ적억제솔대우작담선알,9개신화합물대hCA Ⅱ적억제솔여작담선알상당.결론 hCA Ⅱ억제제사선기술가용우hCA Ⅱ억제제적대규모사선,위발현고효、고아형선택성적탄산항매파향성약물전정기출.진일보심입연구저19개화합물유망획득고효、저부작용적급성고원병방치약물.
Aim 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 source of hCA Ⅱ 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.