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Engineered reversal of the β -oxidation cycle for the synthesis of fuels and chemicals

Engineered reversal of the β -oxidation cycle for the synthesis of fuels and chemicals. 报告人：张文倩 时间： 2011.10.07. Functional reversal of the β -oxidation cycle. 前期准备：对 β -oxidation 的正调控. RB01: MG1655. fadR atoC(c) crp* Δ arcA. 脂肪酸底物不存在的情况下反应仍能进行. 去除 arcA 介导

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Engineered reversal of the β -oxidation cycle for the synthesis of fuels and chemicals

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  1. Engineered reversal of the β-oxidation cycle for thesynthesis of fuels and chemicals 报告人：张文倩 时间： 2011.10.07

  2. Functional reversal of the β-oxidation cycle

  3. 前期准备：对β-oxidation的正调控 RB01: MG1655 fadR atoC(c) crp* ΔarcA 脂肪酸底物不存在的情况下反应仍能进行 去除arcA介导 对β-氧化的抑制作用 使之独立于cAMP RB02: MG1655 fadR atoC(c) crp* ΔarcA ΔadhEΔptaΔfrdA: 封锁其他代谢途径 产乙醇 产乙酸 产琥珀酸

  4. 针对一轮反应，以n-butanol的合成为目标，调节各步反应的基因针对一轮反应，以n-butanol的合成为目标，调节各步反应的基因 • 终止反应： YqhD FucO两个基因分别过表达 Butanol 产量升高 • 第一步 ： acetyl-CoA to acetoacetyl-CoA [atoB+] and [yqeF+] Butanol 产量升高 • 副产物ethanol的产量减少： ΔeutE ΔYqhD

  5. 总结各步反应 （1） YqeF （2） FadB （3） FadB （4） YdiO （5） MhpF and FucO

  6. 第四步：β-氧化中唯一不可逆反应 YdiO FadE ④ • Ydio 替换 FadE A new Escherichia coli metabolic competency: growth on fatty acids by a novel anaerobic β-oxidation pathway. Mol.Microbiol. 47, 793–805 (2003). • 热力学可行性分析： YdiQRST 编码 ferredoxin and flavoproteins ferredoxin 作 enoyl-CoA to acyl-CoA 的还原力ΔG﹤0 • 结论： Ydio –YdiQRST 用于enoyl-CoA to acyl-CoA此步还原反应 逆反应 ③ FadB FadB ② ① YqeF

  7. 最后，与其他合成途径做对比 Malonyl-ACP的合成消耗 ATP acyl-ACP intermediates消耗ATP fatty-acid biosynthesis a nonnatural metabolic engineering approach 目前合成最长碳链水平：己醇 50% C-mole keto-acid pathways 高效性 reversal of the β-oxidation cycle 66.7%C-molebasis 每加两个C原子单元，生成一个ATP 直接以acetyl-CoA作碳原子供体 acyl-CoA intermediates作为n-alcohols和其他产物的前体

  8. Thank you!

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