Although engineered Saccharomyces cerevisiae fermenting cellobiose is useful for the production of biofuels from cellulosic biomass, cellodextrin accumulation is one of the main problems reducing ethanol yield and productivity in cellobiose fermentation with S. cerevisiae expressing cellodextrin transporter (CDT) and intracellular β-glucosidase (GH1-1). In this study, we investigated the reason for the cellodextrin accumulation and how to alleviate its formation during cellobiose fermentation using engineered S. cerevisiae fermenting cellobiose. From the series of cellobiose fermentation using S. cerevisiae expressing only GH1-1 under several culture conditions, it was discovered that small amounts of GH1-1 were secreted and cellodextrin was generated through trans-glycosylation activity of the secreted GH1-1. As GH1-1 does not have a secretion signal peptide, non-conventional protein secretion might facilitate the secretion of GH1-1. In cellobiose fermentations with S. cerevisiae expressing only GH1-1, knockout of TLG2 gene involved in non-conventional protein secretion pathway significantly delayed cellodextrin formation by reducing the secretion of GH1-1 by more than 50%. However, in cellobiose fermentations with S. cerevisiae expressing both GH1-1 and CDT-1, TLG2 knockout did not show a significant effect on cellodextrin formation, although secretion of GH1-1 was reduced by more than 40%. These results suggest that the development of new intracellular β-glucosidase, not influenced by non-conventional protein secretion, is required for better cellobiose fermentation performances of engineered S. cerevisiae fermenting cellobiose.

中文翻译：

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观察到纤维糊精的积累是由工程化的酿酒酵母发酵纤维二糖引起的细胞内 β-葡萄糖苷酶的非常规分泌引起的。

尽管工程化的酿酒酵母发酵纤维二糖可用于从纤维素生物质生产生物燃料，但纤维糊精的积累是使用表达纤维糊精转运蛋白 (CDT) 和细胞内 β-葡萄糖苷酶 (GH1)的酿酒酵母发酵纤维二糖时降低乙醇产量和生产率的主要问题之一-1). 在这项研究中，我们研究了纤维糊精积累的原因，以及如何使用工程化的酿酒酵母发酵纤维二糖来减少纤维二糖发酵过程中纤维糊精的形成。来自使用酿酒酵母的纤维二糖发酵系列在几种培养条件下仅表达 GH1-1，发现分泌少量 GH1-1，并通过分泌的 GH1-1 的转糖基化活性产生纤维糊精。由于 GH1-1 没有分泌信号肽，非常规蛋白分泌可能促进 GH1-1 的分泌。在使用仅表达 GH1-1 的酿酒酵母进行的纤维二糖发酵中，敲除参与非常规蛋白质分泌途径的TLG2基因可通过将 GH1-1 的分泌减少 50% 以上来显着延迟纤维糊精的形成。然而，在用同时表达 GH1-1 和 CDT-1 的酿酒酵母进行的纤维二糖发酵中， TLG2敲除对纤维糊精的形成没有显着影响，尽管 GH1-1 的分泌减少了 40% 以上。这些结果表明，需要开发不受非常规蛋白质分泌影响的新型细胞内 β-葡萄糖苷酶，以提高工程化酿酒酵母发酵纤维二糖的纤维二糖发酵性能。