本文選題：甲醇 + 核酮糖單磷酸途徑��； 參考：《生物技術(shù)通報(bào)》2017年09期

【摘要】：甲醇是一種重要的有機(jī)化工原料,價(jià)格低廉,碳還原度高,在生物化工中是糖質(zhì)原料的理想替代原料。但是常用的工業(yè)微生物宿主如大腸桿菌不能利用甲醇作為碳源,這限制了甲醇在生物化工領(lǐng)域的應(yīng)用。通過表達(dá)甲醇脫氫酶、3-己酮糖-6-磷酸合成酶和6-磷酸-3-己酮糖異構(gòu)酶在大腸桿菌中構(gòu)建可同化甲醇的核酮糖單磷酸途徑。以基因工程菌作為出發(fā)菌株,通過連續(xù)傳代和定向進(jìn)化引入隨機(jī)突變,突變株進(jìn)行以甲醇為碳源的壓力篩選,得到了2株可以利用甲醇生長(zhǎng)的突變株。在以甲醇為輔助碳源的培養(yǎng)基中,25113Δfrm A/p ZWM1-13號(hào)突變株較原始菌株25113Δfrm A/p ZWM1菌體生長(zhǎng)量增加27.6%。對(duì)25113Δfrm A/p ZWM1-13號(hào)突變株進(jìn)行~(13)C示蹤分析檢測(cè)蛋白質(zhì)合成氨基酸,結(jié)果表明氨基酸中~(13)C比例有明顯提高。其中,甲硫氨酸~(13)C標(biāo)記含量增加7.236%。因此,定向進(jìn)化有效地提高了基因工程大腸桿菌的甲醇利用效率。
[Abstract]:Methanol is an important organic chemical raw material with low price and high carbon reduction degree. It is an ideal substitute for sugar raw material in biochemical industry. However, common industrial microbial hosts such as Escherichia coli can not use methanol as a carbon source, which limits the application of methanol in the field of biological and chemical industry. Methanol dehydrogenase 3-hexose 6-phosphate synthase and 6-phosphate -3-hexose isomerase were expressed in Escherichia coli to construct a ribonose monophosphate pathway which could assimilate methanol. Using genetically engineered bacteria as the starting strain, two mutants which can grow with methanol were obtained by continuous passage and directed evolution. The mutants were screened under the pressure of methanol as carbon source. Compared with the original strain 25113 螖 frm A / p ZWM1, the growth of 25113 螖 frm A / p ZWM1 was increased by 27.6% in methanol assisted carbon source medium. The protein synthetic amino acids of 25113 螖 frm A / p ZWM1-13 strain were detected by Tracer analysis. The results showed that the ratio of C13 C in amino acids was significantly increased. The content of methionine 13 C was increased by 7.236%. Therefore, directed evolution can effectively improve the methanol utilization efficiency of genetically engineered Escherichia coli.
【作者單位】： 天津科技大學(xué)生物工程學(xué)院;中國科學(xué)院系統(tǒng)微生物工程重點(diǎn)實(shí)驗(yàn)室;中國科學(xué)院天津工業(yè)生物技術(shù)研究所;
【基金】：國家自然科學(xué)基金青年項(xiàng)目(31700044) 中國科學(xué)院重點(diǎn)部署研究項(xiàng)目(ZDRW-ZS-2016-2)
【分類號(hào)】：Q789
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本文編號(hào)：1918602