重组PDOR基因的表达与纯化及部分酶学性质

Expression,Purification and Characterization of 1,3-Propanediol Oxidoreductase from Klebsiella pneumoniae

将dhaT在E.coliBL21(DE3)pLysS中进行表达,并对含有His6标记的1,3-丙二醇氧化还原酶(PDOR)进行纯化.重组PDOR反应的最适pH值为10.0,最适温度为55℃,其以1,3-丙二醇和NAD+为底物的表观米氏常数Km值分别是15.5,0.23 mmol.L-1.实验表明,Ca2+,Mg2+和Cu2+对酶活性有抑制作用,而Fe2+,Na+,NH4+和Mn2+对酶活性有促进作用;1,3-丙二醇是PDOR适合的氧化底物.

The dhaT gene was expressed with protein in E. coli BL21（DE3）pLysS and the His6-tagged 1,3-propanediol oxidoreductase （PD（）R） was purified. Optimal pH and temperature for purified enzyme were 10.0 and 55 ℃ respectively. The apparent Km values of the enzyme for 1, 3-propanediol and NAD^＋ were 13.5 and 0. 25 mmol · L^-1, respectively. The enzyme could be inhibited by metal ions including Ca^2＋ , Mg^2＋ and Cu^2＋ , while increased by Fe^2＋ Na^＋ , NH4^＋ and Mn^2＋ PDOR showed high specificity for the alcohol substrate.