Raeder Inger Lin Uttakleiv, Moe Elin, Willassen Nils Peder, Smalås Arne O, Leiros Ingar
Department of Chemistry, Faculty of Science, University of Tromsø, N-9037 Tromsø, Norway.
Acta Crystallogr Sect F Struct Biol Cryst Commun. 2010 Feb 1;66(Pt 2):130-6. doi: 10.1107/S1744309109052063. Epub 2010 Jan 26.
The crystal structure of Vibrio cholerae uracil-DNA N-glycosylase (vcUNG) has been determined to 1.5 A resolution. Based on this structure, a homology model of Aliivibrio salmonicida uracil-DNA N-glycosylase (asUNG) was built. A previous study demonstrated that asUNG possesses typical cold-adapted features compared with vcUNG, such as a higher catalytic efficiency owing to increased substrate affinity. Specific amino-acid substitutions in asUNG were suggested to be responsible for the increased substrate affinity and the elevated catalytic efficiency by increasing the positive surface charge in the DNA-binding region. The temperature adaptation of these enzymes has been investigated using structural and mutational analyses, in which mutations of vcUNG demonstrated an increased substrate affinity that more resembled that of asUNG. Visualization of surface potentials revealed a more positive potential for asUNG compared with vcUNG; a modelled double mutant of vcUNG had a potential around the substrate-binding region that was more like that of asUNG, thus rationalizing the results obtained from the kinetic studies.
霍乱弧菌尿嘧啶-DNA N-糖基化酶（vcUNG）的晶体结构已确定至1.5埃分辨率。基于此结构，构建了杀鲑气单胞菌尿嘧啶-DNA N-糖基化酶（asUNG）的同源模型。先前的一项研究表明，与vcUNG相比，asUNG具有典型的冷适应特征，例如由于底物亲和力增加而具有更高的催化效率。asUNG中特定的氨基酸取代被认为通过增加DNA结合区域的正表面电荷而导致底物亲和力增加和催化效率提高。已使用结构和突变分析研究了这些酶的温度适应性，其中vcUNG的突变显示出增加的底物亲和力，更类似于asUNG。表面电位可视化显示，与vcUNG相比，asUNG具有更正的电位；vcUNG的模拟双突变体在底物结合区域周围具有更类似于asUNG的电位，从而使动力学研究获得的结果合理化。
