本文選題：磷酸甘露糖異構(gòu)酶(PMI) + 蠟狀芽孢桿菌��； 參考：《食品工業(yè)科技》2017年06期

【摘要】：將蠟狀芽孢桿菌CZ中的磷酸甘露糖異構(gòu)酶基因(pmi)進(jìn)行克隆,并在大腸桿菌中進(jìn)行異源表達(dá)。將PCR擴(kuò)增得到的pmi基因與p ET-22b(+)表達(dá)載體進(jìn)行連接,轉(zhuǎn)入大腸桿菌BL21(DE3)中,構(gòu)建重組菌株BL21-p ET22b(+)-pmi,并成功表達(dá)了重組磷酸甘露糖異構(gòu)酶。結(jié)果顯示:克隆得到pmi基因序列全長(zhǎng)為948 bp,編碼315個(gè)氨基酸。通過鎳柱His Trap HP親和層析法純化得到具有活性的重組酶,其蛋白分子大小約為40.8 ku。酶學(xué)性質(zhì)結(jié)果顯示:該酶的最適反應(yīng)溫度為35℃,在30~40℃酶活力相對(duì)穩(wěn)定;最適p H為7.0,在弱堿性條件下保存12 h后仍存有50%以上酶活力;不同低濃度的金屬離子Ni~(2+)、Ca~(2+)、Zn~(2+)、Cu~(2+)和Mg~(2+)均對(duì)該酶表現(xiàn)出不同程度的激活作用,其中Mn~(2+)對(duì)該酶激活作用最顯著,當(dāng)其濃度為1 mmol/L時(shí),激活作用最大,而Co~(2+)對(duì)其有明顯的抑制作用。
[Abstract]:The phosphomannose isomerase gene of Bacillus cereus CZ was cloned and expressed in E. coli. The pmi gene amplified by PCR was ligated with pET-22b () expression vector and transferred into E. coli BL21DE3. The recombinant strain BL21-p ET22b (-pmib) was constructed, and the recombinant phosphate mannose isomerase was successfully expressed. The results showed that the total length of pmi gene was 948 BP, encoding 315 amino acids. The recombinant enzyme was purified by nickel column His Trap HP affinity chromatography and its protein molecular size was about 40. 8 ku. The results of enzymatic properties showed that the optimum reaction temperature was 35 鈩，

本文編號(hào)：1946543