Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, 100081, Beijing, People's Republic of China.
Mol Biol Rep. 2011 Apr;38(4):2437-43. doi: 10.1007/s11033-010-0379-5. Epub 2010 Nov 17.
The -1 subsite of bacterial fructansucrases (FSs) (levansucrases and inulosucrases) plays an important role in the substrate recognition, binding and catalysis. Three residues (for example W47, W118 and R193, Zymomonas mobilis levansucrase numbering) at the -1 subsite are completely conserved among FSs. Site-directed mutational analysis showed that the substitutions of the three strictly conserved amino acid residues, W47N, W47H, W118N, W118H, R193K and R193H, significantly decreased enzyme activities and synthesis rates of levan, while the size of the synthesized oligosaccharides had been influenced. These experimental results, combined with 3D structure modeling, lead to our proposal that a single amino acid residue change in subsite -1 of levansucrase can influence change to the size and polarity of the sucrose binding pocket with a concomitant change to substrate binding and catalysis, and thus having an overall influence on the enzyme activities and products.
细菌果聚糖蔗糖酶（FSs）（蔗聚糖酶和菊粉蔗糖酶）的-1 亚位在底物识别、结合和催化中起着重要作用。FSs 中的-1 亚位完全保守三个残基（例如，Zymomonas mobilis 蔗聚糖酶编号中的 W47、W118 和 R193）。定点突变分析表明，三个严格保守的氨基酸残基 W47N、W47H、W118N、W118H、R193K 和 R193H 的取代显著降低了酶活性和蔗聚糖的合成速率，而合成寡糖的大小也受到了影响。这些实验结果与 3D 结构建模相结合，使我们提出这样的假设，即-1 亚位的单个氨基酸残基变化可以影响蔗糖结合口袋的大小和极性的变化，进而影响底物结合和催化，从而对酶活性和产物产生整体影响。
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