毛白杨4-香豆酸:辅酶A连接酶338Val缺失突变分析

通过对毛白杨中的4--香豆酸:辅酶A连接酶1（Pt4CL1）蛋白第338位缬氨酸进行缺失突变，从而获得了具有芥子酸催化活性的Pt4CL1蛋白突变体338dVal。与野生型4CL1蛋白活性相比较，该突变体获得了催化芥子酸的活性，比活力是(162±034) nkat/mg。同时，对4--香豆酸催化活性有轻微的降低（约降低12％），对咖啡酸的催化活性有明显增加（约增加126％），对阿魏酸的催化活性有部分的降低（约降低29％），对肉桂酸的催化活性没有显著变化。该结论证实了第338位缬氨酸对底物芥子酸5位甲氧基具有空间位阻效应。该项研究为通过基因工程技术调控木质素的合成提供了新的方法。 

Catalytic properties of 338Val-deleted mutant of 4-coumarate:coenzyme A ligase from Populus tomentosa.

4-coumarate:CoA ligase 1 (4CL1) is a key rate-limiting enzyme in the pathway of lignin biosynthesis. Studying the catalytic properties of 4CL1 enzyme is useful for understanding the pathway of lignin biosynthesis. This paper reports that a Pt4CL1 variant 338dVal, which is a deleted Val-338 in amino acid sequence of 4-coumarate:CoA ligase from Populus tomentosa, gains the function of using sinapic acid. The specific activity to sinapic acid was (1.62±0.34) nkat/mg. Meanwhile, the catalytic activity of 338dVal to 4-coumarate acid decreased by 12% approximately; the catalytic activity of 338dVal to caffeic acid increased by 126% and the catalytic activity of 338dVal to ferulic acid decreased by 29% roughly. The catalytic acitivity of 338dVal to cinnamic acid did not change. The results show that the Val-338 in Pt4CL1 has size exclusion effect on 5-methoxy of sinapic acid. The paper provides a new method that can be used to regulate the lignin biosynthesis using genetic engineering.