中文摘要：

小麦种子过氧化物酶WPl属于含血红素的植物III型过氧化物酶，该酶不仅具有抗真菌活性，而且影响面粉加工品质。为提高WPl在大肠杆菌中的功能性表达，构建了用于提高大肠杆菌内源血红素合成，包含hemA和hemL基因的重组质粒pACYC．A—L，将其分别与包含WPI基因的分泌型和非分泌型表达载体pMAL．p4x—WPl与pET2la—MBP-WPl共同转化大肠杆菌T7Express菌株；利用直链淀粉（Amylose）亲和层析柱纯化获得MBP．WPl融合蛋白，并以2，2’一联氮．二（3．乙基苯并噻唑．6．磺酸）二铵盐（ABTS）为底物检测重组WPl的催化能力。结果表明，含pACYC—A．L的宿主菌28℃诱导12h后，培养液中5。氨基乙酰丙酸（5-ALA）含量可达146．73mg／L，卟啉类物质含量也显著上升。共转化pACYC-A—L和pMAL—p4x-WPl比单独转化pET21a。MBP．WP】获得的重组WPl的比活力提高14．6倍。该研究不仅成功地增强了小麦WPl在大肠杆菌中的功能性表达，同时为其他具有重要生物学功能并含血红素辅基蛋白的功能性表达提供了有益参考。

英文摘要：

Wheat grain peroxidase 1 （WP1） belonged to class III plant peroxidase with cofactor heme, which not only has antifungal activity, but also influences the processing quality of flour. In order to enhance functional expression of WP1 in prokaryotic system by increasing endogenous heme synthesis, we constructed a recombinant plasmid pACYC-A-L containing hemA and hemL of Esherichia coli. Then, we co-transformed it into host strain T7 Express with secretive expression vector （pMAL-p4x-WP 1） or non-secretive expression vector （pET2 la-MBP-WP 1）, respectively. The MBP-WP 1 fusion protein was further purified by amylose affinity chromatography and its peroxidase activity was assayed using 2,2＇-azino-bis （3-ethylbenzothiazoline-6-sulfonate） （ABTS） as substrate. At 12 h after induction at 28 degree, the extracellular 5-aminolevulinic acid （5-ALA） production of T7 Express/pACYC-A-L was up to 146.73 mg/L, simultaneously the extracellular porphrins also increased dramatically. The peroxidase activity of functional MBP-WP1 obtained from T7 Express/ （pACYC-A-L ＋ pMAL-p4x-WP1） was 14.6-folds of that purified from T7 Express/ pET21a-MBP-WP1. This study not only successfully enhanced functional expression of wheat peroxidase 1 in Esherichia coli, but also provided beneficial references for other important proteins with cofactor heme.