Development of anti-Trypanosoma drug based on structural and functional analysis of novel L-amino acid dehydrogenase

利用重叠引物扩增了布氏锥虫L-苏氨酸脱氢酶（ThrDH）基因，并将其与表达载体pCold TF连接。 然后用载体转化大肠杆菌，并通过向培养基中加入1.0 mM IPTG进行诱导。 该基因在E. coli，并对产物进行了纯化和表征。 SDS-PAGE显示T.结果表明，该蛋白的分子量约为81 kDa，与从氨基酸序列计算的分子量一致。 测定轻微NAD依赖性ThrDH活性。

L-Threonine dehydrogenase (ThrDH) gene from the pathogenic parasite Trypanosoma brucei was amplified using the overlapping primers, and ligated with the expression vector pCold TF. Escherichia coli was then transformed with the vector and induced by adding 1.0 mM IPTG to the medium. The gene was overexpressed in E. coli, and its product was purified and characterized. SDS-PAGE showed the subunit molecular mass of T. brucei ThrDH to be about 81 kDa, which was consistent with the molecular weight calculated from the amino acid sequence. The slightly NAD-dependent ThrDH activity was determined.