Synergistically enhanced thermostability of Saccharomyces cerevisiaeby ubiquitin-like protein mediation and heat shock response

doi:10.11949/j.issn.0438-1157.20151778

Abstract

Abstract:

To improve the thermostability and fermentation performance of Saccharomyces cerevisiae to reduce the energy consumption of the cooling progress in industrial fermentation, the protein homeostasis was regulated through ubiquitin-like protein mediation and heat shock response. In this study, many heat-resistant gene devices were mined out from genes related to protein homeostasis and constructed with the regulatory device FBA1p, and then transformed into Saccharomyces cerevisiae INVSC1. Outstanding heat-resistant devices FBA1p-atg8 and FBA1p-hsp104 were screened through gradually increased temperature incubation. Compared with the control, the OD₆₆₀ of the engineered yeast strains S.c-ATG8 and S.c-HSP104 were both over 50% higher (84 h) and their cell viability were 1.64 to 3.01 times higher (72 h) when cultured at 40℃. The physiological characteristics implied that the thermotolerant strains possessed better cell wall integrity and higher trehalose content. In order to strengthening the regulatory mechanisms of both ubiquitin-proteasome system pathway and heat-shock responses within the network of protein homeostasis, atg8 and hsp104 were assembled to construct bifunctional engineered strain S.c-ATG8-HSP104, which showed better growth ability, stronger cell activity and higher ethanol yield at 40℃. The results revealed that the synergistic effect of ubiquitin-like protein and heat shock protein could enhance yeast thermotolerance and improve strain activity.

Key words: synthetic biology, fermentation, biotechnology, ubiquitin-like protein, heat shock protein, protein homeostasis, thermostability

摘要：

通过调节类泛素和热激响应介导的酿酒酵母内部的活性蛋白质平衡,提高酵母细胞的热稳定性和乙醇发酵性能,从而达到工业生产中降低控温能耗的目的。将5个蛋白质平衡相关基因分别与调控基因FBA1p组合构建耐热元器件,并将其导入酿酒酵母Saccharomyces cerevisiaeINVSC1,通过梯度升温培养筛选得到能赋予酵母细胞较好耐热性的类泛素元器件FBA1p-atg8和热激蛋白元器件FBA1p-hsp104;其对应的工程菌S.c-ATG8和S.c-HSP104在40℃恒定培养下,OD₆₆₀值均比对照高50%以上(84h),细胞存活率分别是对照的1.64倍和3.01倍(72h),且都具有较好的细胞壁完整性及海藻糖合成量。将atg8与hsp104组合构建成双功能耐热元器件,其工程菌S.c-ATG8-HSP104在40℃恒定发酵的生长能力、细胞活力和乙醇生产能力都明显优于S.c-ATG8与S.c-HSP104。结果表明,通过类泛素介导与热激蛋白响应协同调节胞内活性蛋白质平衡可以有效地提高酿酒酵母的热稳定性。

关键词: 合成生物学, 发酵, 生物技术, 类泛素, 热激蛋白, 蛋白质平衡, 热稳定性

CLC Number:

TQ028.8

XIAO Bing, LI Jun, LI Chun. Synergistically enhanced thermostability of Saccharomyces cerevisiaeby ubiquitin-like protein mediation and heat shock response[J]. CIESC Journal, 2016, 67(6): 2503-2509.

肖冰, 李珺, 李春. 类泛素介导和热激响应协同提高酿酒酵母的热稳定性[J]. 化工学报, 2016, 67(6): 2503-2509.

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