北京林业大学学报2016，Vol.38Issue(7)：9-15,7.DOI:10.13332/j.1000-1522.20160121
毛果杨中链酰基辅酶A合成酶的克隆及酶学分析
Cloning and enzymatic analysis of medium-chain acyl coenzyme A synthetase in Populus trichocarpa
摘要
Abstract
Medium-chain acyl coenzyme A synthetase ( MACS ) family is a subfamily of adenylate-forming enzymes superfamily, catalyzing the medium-chain fatty acids with CoA to produce medium-chain-acyl-CoA. PtMACS1 (gene model:estExt fgenesh4 pg. c 640066) was cloned via blast in the database JGI of Populus trichocarpa. Sequence analysis showed that the conserved domains BoxI and BoxII in 4CL were not conserved in the proteins. Expression vector PtMACS1-pET-30a ( + ) was constructed and transformed into E. coli BL21 (DE3) to express the recombinant protein. The recombinant protein was purified by Ni-NTA affinity chromatography, enzymatic analysis showed that the recombinant protein had remarkable catalytic activity to the medium-chain fatty acids, such as hexanoic acid, nonanoic acid and decylic acid, and the Kcat were 130, 193 and 201μmol/L/(min·mg) respectively. When decylic acid was used as the substrate, the optimum temperature was 37℃ and pH was 7. 0. The results demonstrate that PtMACS1 is one of the MACS family members, and supplies research data in identifying and classifying members from adenylate-forming enzymes superfamily in Populus trichocarpa.关键词
毛果杨/MACS/克隆/原核表达/酶学分析Key words
Populus trichocarpa/MACS/gene clone/prokaryotic expression/enzymatic analysis分类
农业科技引用本文复制引用
曹山,蒋璐瑶,李丽红,要笑云,张强,撖静宜,王莹,李慧,陆海..毛果杨中链酰基辅酶A合成酶的克隆及酶学分析[J].北京林业大学学报,2016,38(7):9-15,7.基金项目
国家自然科学基金项目(31570582、30671697、J1103516)。 （31570582、30671697、J1103516）
