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ESTABLISHMENT AND APPLICATION OF HIGH THROUGHPUT SCREENING MODEL FOR INFLUENZA VIRUS NEURAMINIDASE INHIBITORS IN VITRO
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- Author:
- No author available
- Journal Title:
- ACTA PHARMACEUTICA SINICA
- Issue:
- 12
- DOI:
- 10.3321/j.issn:0513-4870.2002.12.005
- Key Word:
- 神经氨酸酶;甲、乙型流感病毒;筛选模型
Abstract： 目的建立适用于高通量筛选的流感病毒神经氨酸酶(neuraminidase,NA)抑制剂筛选模型.方法从甲型及乙型流感病毒中制备神经氨酸酶,以2′-(4-methylumbelliferyl)-α-D-N-acetylneuraminic acid(MUNANA)作为底物,建立检测神经氨酸酶活性的荧光测定法及其抑制剂体外筛选方法,用高通量筛选系统对1 200个化合物与提取物进行初筛.结果神经氨酸酶酶促反应以pH 3.5,二价阳离子浓度为2～6 mmol*L-1及37℃孵育时酶活性最佳;甲、乙型流感病毒不同株神经氨酸酶的米氏常数(Km)的范围为(4.89～5.94) μmol*L-1;初筛发现12个化合物对流感病毒神经氨酸酶有可重复的抑制活性.结论优化了神经氨酸酶反应体系,建立的体外模型可用于抗甲、乙型流感病毒药物的高通量筛选及酶抑制动力学的研究.
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