Engineering endogenous ABC transporter with improving ATP supply and membrane flexibility enhances the secretion of β-carotene in Saccharomyces cerevisiae.
Engineering endogenous ABC transporter with improving ATP supply and membrane flexibility enhances the secretion of β-carotene in Saccharomyces cerevisiae.
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通过改善 ATP 供应和膜灵活性改造内源 ABC 转运蛋白可增强酿酒酵母中 β-胡萝卜素的分泌
DOI：
10.1186/s13068-020-01809-6
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发表时间：
2020
影响因子：
6.3
通讯作者：
Yan GL
中科院分区：
文献类型：
--
作者：
Bu X;Lin JY;Cheng J;Yang D;Duan CQ;Koffas M;Yan GL
产品毒性是微生物生产生物燃料的瓶颈之一，转运体介导的生物燃料分泌为解决这一问题提供了一个有希望的策略。作为工业规模生产生物燃料的强大微生物宿主，酿酒酵母菌含有强大的运输系统，可以出口各种有毒化合物以维持生存。本研究的目的是利用内源性ABC转运蛋白结合生理工程，改善酿酒酵母疏水产物β-胡萝卜素的分泌和产生。底物诱导性是大多数内源性转运蛋白的一个显著特征。通过比较蛋白质组学分析和转录确认，我们确定了5种潜在的ABC转运蛋白（Pdr5p、Pdr10p、Snq2p、Yor1p和Yol075cp）参与β-胡萝卜素外排。β-胡萝卜素的积累还从能量代谢、线粒体翻译、脂质代谢、麦角甾醇生物合成过程、细胞壁合成等多方面影响细胞生理。本研究采用诱导型GAL启动子过表达候选转运蛋白，增强了β-胡萝卜素的分泌和胞内生成，其中Snq2p表现最好，分别比亲本菌株YBX-01提高了4.04倍和1.33倍。为了进一步提高外排能力，采用了增加ATP供给和提高膜流动性两种策略。与亲本菌株YBX-01相比，工程菌株YBX-20的β-胡萝卜素分泌量增加了5.80倍，细胞内β-胡萝卜素产量增加了1.71倍。总之，我们的研究结果表明，设计内源性质膜ABC转运蛋白是酿酒酵母疏水产物外排的一种很有前途的方法。我们还强调了改善细胞生理以提高ABC转运体效率的重要性，特别是能量状态和细胞膜性质。
Product toxicity is one of the bottlenecks for microbial production of biofuels, and transporter-mediated biofuel secretion offers a promising strategy to solve this problem. As a robust microbial host for industrial-scale production of biofuels, Saccharomyces cerevisiae contains a powerful transport system to export a wide range of toxic compounds to sustain survival. The aim of this study is to improve the secretion and production of the hydrophobic product (β-carotene) by harnessing endogenous ABC transporters combined with physiological engineering in S. cerevisiae. Substrate inducibility is a prominent characteristic of most endogenous transporters. Through comparative proteomic analysis and transcriptional confirmation, we identified five potential ABC transporters (Pdr5p, Pdr10p, Snq2p, Yor1p, and Yol075cp) for β-carotene efflux. The accumulation of β-carotene also affects cell physiology in various aspects, including energy metabolism, mitochondrial translation, lipid metabolism, ergosterol biosynthetic process, and cell wall synthesis. Here, we adopted an inducible GAL promoter to overexpress candidate transporters and enhanced the secretion and intracellular production of β-carotene, in which Snq2p showed the best performance (a 4.04-fold and a 1.33-fold increase compared with its parental strain YBX-01, respectively). To further promote efflux capacity, two strategies of increasing ATP supply and improving membrane fluidity were following adopted. A 5.80-fold increase of β-carotene secretion and a 1.71-fold increase of the intracellular β-carotene production were consequently achieved in the engineered strain YBX-20 compared with the parental strain YBX-01. Overall, our results showcase that engineering endogenous plasma membrane ABC transporters is a promising approach for hydrophobic product efflux in S. cerevisiae. We also highlight the importance of improving cell physiology to enhance the efficiency of ABC transporters, especially energy status and cell membrane properties.
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影响因子：
14.8
作者：
Gietz, R. Daniel;Schiestl, Robert H.
通讯作者：
Schiestl, Robert H.
DOI：
10.1073/pnas.1301358110
发表时间：
2013-05-07
影响因子：
11.1
作者：
Doshi, Rupak;Tuan Nguyen;Chang, Geoffrey
通讯作者：
Chang, Geoffrey
影响因子：
3.5
作者：
Claus, Silke;Jezierska, Sylwia;Van Bogaert, Inge N. A.
通讯作者：
Van Bogaert, Inge N. A.
影响因子：
3.9
作者：
HAGGERTY, DF;KALRA, VK;CHIAPPELLI, F
通讯作者：
CHIAPPELLI, F
影响因子：
4.4
作者：
Li, Y;Chen, J;Koo, YM
通讯作者：
Koo, YM