Transketolase is the key enzyme of photosynthesis pathways. It is of great importance for herbicide seeking through new herbicidal substance targeting transketolase. In this study, homologous protein of transke-tolase was obtained through Modeler and optimized using Amber12 , its binding sites of pivotal amino acids were identified with bioinformatics tools. Interaction between AtTKL1 and α-terthienyl finally got verified by spectroscopy and enzyme activity analysis. The three dimensional frame model of transketolase protein AtTKL1 in Arabidopsis thaliana was homologically modeled according to Maize transketolase protein 1ITZ, revised by dynamic molecular methods. With bioinformatics tools, the sites in AtTKL1 were determined, His143, Gly234, Asn263, Arg434, Ser461, Gln488, Phe515, His539, Asp547and Arg598 for binding, His103 and His340 for catalysis. The molecular docking of AtTKL1 and α-terthienyl was performed by computer simula-tion, and the results confirmed the perfect match ofα-terthienyl with active cleavage of AtTKL1. The binding sites His143 and Gly234 , and the catalytic sites His103 and His340 were the responsible binding sites ofα-terthienyl. Prokaryotic expression of the protein and fluorescence quenching spectroscopy have verified the combination of α-terthienyl and AtTKL1 . The reduction on the activity of transketolase treatmented withα-terthienyl gave a further confirmation about the interaction between AtTKL1 and α-terthienyl. The results will contribute to exploiting new efficient and low toxic herbicides.%以玉米转酮醇酶(1ITZ)的晶体结构为模板,对拟南芥转酮醇酶(AtTKL1)的三维结构进行模拟和优化.利用生物信息学方法,确定其氨基酸结合位点为 His143, Gly234, Asn263, Arg434, Ser461, Gln488, Phe515, His539, Asp547和Arg598,催化氨基酸位点为His103和His340.通过分子模拟确定α-三联噻吩与AtTKL1的活性口袋可有效结合,且AtTKL1的催化位点和结合位点为其与α-三联噻吩结合的关键位点.利用荧光猝灭光谱技术确定AtTKL1与α-三联噻吩之间存在结合作用；酶活力检测结果表明加入α-三联噻吩后转酮醇酶蛋白的活性降低,进一步验证了AtTKL1与α-三联噻吩之间的相互作用.
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