McKetta Department of Chemical Engineering, The University of Texas at Austin, 200 East Dean Keeton St., Stop C0400, Austin, TX, 78712, USA.
Institute for Cellular and Molecular Biology, The University of Texas at Austin, 2500 Speedway Avenue, Austin, TX, 78712, USA.
J Ind Microbiol Biotechnol. 2019 Dec;46(12):1715-1724. doi: 10.1007/s10295-019-02228-9. Epub 2019 Aug 19.
Ionic liquids show promise for deconstruction of lignocellulosic biomass prior to fermentation. Yet, imidazolium ionic liquids (IILs) can be toxic to microbes even at concentrations present after recovery. Here, we show that dominant overexpression of an Ilt1p homolog (encoded by YlILT1/YALI0C04884) from the IIL-tolerant yeast Yarrowia lipolytica confers an improvement in 1-ethyl-3-methylimidazolium acetate tolerance in Saccharomyces cerevisiae compared to the endogenous Ilt1p (ScILT1/YDR090C). We subsequently enhance tolerance in S. cerevisiae through directed evolution of YlILT1 using growth-based selection, leading to identification of mutants that grow in up to 3.5% v/v ionic liquid. Lastly, we demonstrate that strains expressing YlILT1 variants demonstrate improved growth rate and ethanol production in the presence of residual IIL. This shows that dominant overexpression of a heterologous protein (wild type or evolved) from an IIL-tolerant yeast can increase tolerance in S. cerevisiae at concentrations relevant to bioethanol production from IIL-treated biomass.
离子液体在木质纤维素生物质发酵前的解构中显示出巨大的潜力。然而，即使在回收后存在的浓度下，咪唑类离子液体（IILs）也可能对微生物有毒。在这里，我们表明，来自耐受 IIL 的酵母解脂耶氏酵母（Yarrowia lipolytica）的 Ilt1p 同源物（由 YlILT1/YALI0C04884 编码）的过度表达，与内源性 Ilt1p（ScILT1/YDR090C）相比，可提高对 1-乙基-3-甲基咪唑𬭩乙酸盐的耐受性。随后，我们通过使用基于生长的选择对 YlILT1 进行定向进化，提高了 S. cerevisiae 的耐受性，从而鉴定出可在高达 3.5%v/v 的离子液体中生长的突变体。最后，我们证明表达 YlILT1 变体的菌株在残留 IIL 存在下表现出更快的生长速度和更高的乙醇产量。这表明，从 IIL 处理的生物质生产生物乙醇的相关浓度下，来自耐受 IIL 的酵母的异源蛋白（野生型或进化型）的过度表达可以提高 S. cerevisiae 的耐受性。
