中文摘要：

将拟南芥Antiquitin基因重组到原核表达载体pMAL-c4x和pET41中,在T7 Express菌株中诱导表达,经Amylose和Ni-NTA亲和层析柱纯化获得重组蛋白。SDS-PAGE结果表明：MBP和His-tag融合的拟南芥An-tiquitin主要以可溶性形式存在,表达量分别占细胞总蛋白的25.1%和39.4%。以乙醛和NAD＋为底物测定融合蛋白活性,结果显示：His-tag融合的Antiquitin具有醛脱氢酶活性,比活力为8.98 U/mg,乙醛的表观Km和Vmax值分别为0.98 mmol/L和12.75 U/mg。序列比对和结构预测结果显示：拟南芥Antiquitin包含该家族蛋白典型的催化结构域、NADH结合结构域和寡聚化结构域,活性中心残基为Gly238、Gly291、Glu391、Phe393。

英文摘要：

The encoding sequence of Antiquitin protein from Arabidopsis thaliana was cloned into pMAL-c4x and pET41 vector.Fusion proteins were expressed in E.coli strain T7 Express and purified by amylose resin column and Ni-NTA resin column.Based on the results of SDS-PAGE,the yield of fusion protein,which tagged by MBP and His6,accounts for 25.1% and 39.4% of the total bacterial protein,respectively.The aldehyde dehydrogenase activity of the purified protein was determined using acetaldehyde and NAD＋ substrates.The result showed that the soluble fusion protein from Ni-NTA resin column had detectable aldehyde dehydrogenase activity and the specific activity is 8.98 U/mg.The Km and Vmax for acetaldehyde are 0.98 mmol/L and 12.75 U/mg,respectively.Sequence alignment and structure prediction reveals：Arabidopsis Antiquitin may exist as an oligomer,each monomer is comprised of three domains,catalytic domain,NAD＋-binding domain and oligomerization domain,and the active residues are Gly238,Gly291,Glu391 and Phe393.