Jiangnan University

Xiaowei Yu

Yan Xu

The invention discloses a high-secretion heat-resistant yeast genetically engineered strain and application thereof, belonging to the field of biotechnology. Mutagenesis and domestication are performed to obtain the yeast genetically engineered strain capable of expressing a lipase gene at a high secretion level at high temperature. The strain is collected by China Center for Type Culture Collection (CCTCC), and the collection number is CCTCC NO: M 2016278. The sequencing analysis shows that the lipase gene and promoter sequence thereof are not mutated, which indicates that the high-secretion expression of the lipase gene by the mutant strain is caused by mutation of other gene sequences in the genome. By knocking out the lipase gene in homologous double-crossover way, the constructed knockout strain can be used as an expression host of other exogenous proteins, can be used for enhancing the expression level of other exogenous genes, and can be subjected to fermentation under high-temperature fermentation conditions.

High-secretion heat-resistant yeast genetically engineered strain and application thereof

The invention discloses a method for improving the extracellular expression level of a foreign protein by means of phospholipase fusion expression. Four proteins, PLA, MBP, CBD and SUMO, are used as a fusion tag to construct a fusion gene. Compared with an original protein MOH without any fusion tag, the extracellular expression level and enzymatic activity of all the four fusion proteins are increased to some degree. Among them, the fusion protein using PLAas the fusion tag has the highest expression level, which is 7.4 times higher than that of the original protein. Compared with other fusion tags, PLAhas a low molecular weight and the fusion protein having PLA2 as the fusion tag has the highest expression level (up to 12 g&#xb7;Lin a 7 L fermentation tank for high-density fermentation). It is shown that the secretory expression of a foreign protein can be effectively increased by using PLAas a fusion tag.

Methods for improving expression levels of foreign proteins by means of phospholipase fusion expression

The present invention provides a novel dual-function lipase mutant and its application in processing of flour products. The amino acid sequence of the lipase has one of the following amino acid substitutions: P298T,P298T/H317P, P298T/H317P/V326S, P298T/T218S/S234F, P298T/H317P/P168L/A129S, P298T/S234F/K161R/V326S, wherein the substitutions are relative to a parent amino acid sequence set forth in SEQ ID NO:1; wherein the lipase mutant maintains a triglyceride and lecithin hydrolysis activity and good performance in thermostability. The mutants have good performance in processing of flour products, while they can significantly whiten the bread or other products in processing of flour products and significantly increase the specific volume in bread baking process.

Bifunctional lipase mutant and methods of using same

The present invention provides a novel dual-function lipase variant and its application in processing of flour products. The amino acid sequence of the lipase has one of the following amino acid substitutions: P298T, P298T/H317P, P298T/H317P/V326S, P298T/T218S/S234F, P298T/H317P/P168L/A129S, P298T/S234F/K161R/V326S or the nucleotide sequence of wherein said lipase is substituted, deleted, or added based on the sequence encoding the amino acid described in (a) and has at least 80% identity with it. The variants have good performance in processing of flour products, while they can significantly whiten the bread or other products in processing of flour products and significantly increase the specific volume in bread baking process.

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Wuxi, China

Xiaowei Yu