玉米大斑病菌支链氨基酸氨基转移酶基因的克隆与原核表达
Gene Cloning and Prokaryotic Expression of Branched-Chain Amino Acid Transaminase in Setosphaeria turcica
张运峰 1张淑红 1许可 1高凤菊 1武秋颖 1石洪凌 1李艳梅 1范永山1
作者信息
- 1. 唐山师范学院生命科学系/唐山市农业病原真菌与毒素重点实验室,河北唐山 063000
- 折叠
摘要
支链氨基酸(Branched chain amino acid,BCAA)是生物体内重要的营养物质和信号分子,对蛋白质合成、糖代谢、氧化应激等有重要的调控作用.支链氨基酸氨基转移酶(branched-chain amino acid transaminase,BCAT)是BCAA分解代谢的关键酶,参与许多生理和病理过程.为明确BCAT对玉米大斑病菌(Setosphaeria turcica)生长发育和侵染的影响,克隆玉米大斑病菌BCAT基因(StBCAT1),该基因编码的StBCAT1含有保守结构域BCAT_beta_fam-ily,属于PLPDE_Ⅳ超家族,与玉米圆斑病菌(Helminthosporium carbonum)产生的寄主选择性毒素HC-toxin ToxF高度同源.为了获得大量高纯度的StBCAT1蛋白,构建该基因的原核表达载体,并转化BL21(DE3)菌株.经IPTG诱导表达,SDS-PAGE和Western Blotting检测,结果表明,StBCAT1基因在大肠杆菌得到高效表达,获得了预期分子量为39.2 KD 的 StBCAT1蛋白.
Abstract
The branched chain amino acid(BCAA)is important nutrient and signaling molecule regulating pro-tein synthesis,sugar metabolism and oxidative stress in living cells.Branched chain amino acid transaminase(BCAT)is the key enzyme in the catabolism of BCAA and involves in many physiological and pathological process-es.In order to clarify the effects of BCAT on the growth,development and infection of Setosphaeria turcica,the StB-CAT1 gene was cloned,whose coding protein contains a conserved domain of BCAT_beta_family belonging to PLP-DE_Ⅳ superfamily,and is highly homologous to host-selective HC-toxin ToxF of Helminthosporium carbonum.With the aim to obtain highly purified StBCAT1 protein,the prokaryotic expression vector of StBCAT1 was con-structed and transformed into bacterial strain BL21(DE3).After induced by IPTG,the StBCAT1 was efficiently ex-pressed in Escherichia coli and the expected protein(about 39 KD)was gotten after detected by SDS-PAGE and Western Blotting.
关键词
玉米/大斑病菌/基因克隆/支链氨基酸转移酶/原核表达Key words
Maize/Setosphaeria turcica/Gene cloning/Branched chain amino acid transaminase/Prokaryotic expression引用本文复制引用
基金项目
河北省重点研发计划项目(21374301D)
河北省自然科学基金项目(B2023105007)
河北省自然科学基金项目(C2023105003)
唐山师范学院重点培育项目(ZD-PY06)
唐山师范学院重点培育项目(ZDPY08)
唐山市科技计划项目(21130201C)
出版年
2024
NETL
NSTL
万方数据