Study of MMLV RT-Binding with DNA using Surface Plasmon Resonance Biosensor

作     者：Lei WU~1 Ming-Hui HUANG~2 Jian-Long ZHAO~(1*) Meng-Su YANG~~(1,2*) 1 Shanghai Institute of Microsystem and Information Technology,Chinese Academy of Sciences,Shanghai 200050,China 2 Department of Bioloy and Chemistry,City University of Hong Kong,Hong Kong,China Lei WU~1 Ming-Hui HUANG~2 Jian-Long ZHAO~(1*) Meng-Su YANG~~(1,2*) 1 Shanghai Institute of Microsystem and Information Technology,Chinese Academy of Sciences,Shanghai 200050,China 2 Department of Bioloy and Chemistry,City University of Hong Kong,Hong Kong,China

作者机构：Shanghai Institute of Microsystem and Information Technology Chinese Academy of SciencesShanghai 200050 China Department of Biology and Chemistry City University of Hong KongHong Kong China 

出 版 物：《Acta Biochimica et Biophysica Sinica》 (生物化学与生物物理学报(英文版))

年 卷 期：2005年第37卷第9期

页      面：634-642页

核心收录：

学科分类：0710[理学-生物学] 07[理学] 071007[理学-遗传学] 

基　　金：This work was supported by a grant from the Major Basic Research Program of the Science and Technology Commission Foundation of Shanghai (No.04JC14081) 

主　　题：surface plasmon resonance biosensor reverse transcriptase kinetics inhibitor 

摘      要：Surface plasmon resonance biosensor technique was used to study the binding of Moloneymurine leukemia virus reverse transcriptase without RNase H domain （MMLV RT-） with DNA in the ab-sence and in the presence of *** DNA substrates,including single-stranded DNA （ssDNA）,DNA template-primer （T-P） duplex and gapped DNA,were immobilized on the biosensor chip surface usingstreptavidin-biotin,and MMLV RT--DNA binding kinetics were analyzed by different *** RT-could bind with ssDNA and the binding was involved in conformation *** RT- binding DNA T-Pduplex and gapped DNA could be analyzed using the simple 1:1 Langmuir *** lack of RNase Hdomain reduced the affinity between MMLV RT- and T-P *** effects of RT inhibitors,includingefavirenz,nevirapine and quercetin,on the interaction between MMLV RT- and gapped DNA were analyzedaccording to recovered kinetics *** slightly interfered with the binding between RT andDNA and the affinity constant in the presence of the inhibitor (KA=1.21×106 M-1) was lower than in theabsence of the inhibitor (KA=4.61×106 M-1).Nevirapine induced relatively tight binding between RT andDNA and the affinity constant in the presence of the inhibitor (KA=1.47×107 M-1) was approximately threefolds higher than without nevirapine,mainly due to rapid association and slow ***,aflavonoid originating from plant which has previously shown strong inhibition of the activity of *** to have minimal effect on the RT-DNA binding.