caa a22 4500 606212841 CHVBK 20210128101044.0 cr unu---uuuuu 210128e20150801xx s 000 0 eng 10.1007/s10295-015-1633-6 doi (NATIONALLICENCE)springer-10.1007/s10295-015-1633-6 Combined use of GAP and AOX1 promoters and optimization of culture conditions to enhance expression of Rhizomucor miehei lipase [Elektronische Daten] [Dong He, Wen Luo, Zhiyuan Wang, Pengmei Lv, Zhenhong Yuan] Rhizo mucor miehei lipase (RML) is an industrially important enzyme, but its application is limited due to its high cost. In this study, a series of measures such as codon optimization, propeptide addition, combined use of GAP and AOX1 promoters, and optimization of culture conditions were employed to increase the expression of RML. Three transformants of the constitutive-inducible combined Pichia pastoris strains were generated by transforming the pGAPZαA-rml vector into the pPIC9K-rml/GS115 strain, which resulted in high-expression yields of RML. Using the shake flask method, highest enzyme activity corresponding to 140 U/mL was observed in the strain 3-17, which was about sixfold higher than that of pPIC9K-rml/GS115 or pGAPZαA-rml/GS115. After optimization of culture conditions by response surface methodology, the lipolytic activity of strain 3-17 reached 175 U/mL in shake flasks. An increase in the copy number simultaneously with the synergistic effect provided by two promoters led to enhanced degree of protein expression. Society for Industrial Microbiology and Biotechnology, 2015 RML nationallicence Pichia pastoris nationallicence P GAP nationallicence P AOX1 nationallicence Optimization nationallicence He Dong Key Laboratory of Renewable Energy, Guangzhou Institute of Energy Conversion, Chinese Academy of Sciences, No. 2, Nengyuan Road, Tianhe Area, 510640, Guangzhou, People's Republic of China aut Luo Wen Key Laboratory of Renewable Energy, Guangzhou Institute of Energy Conversion, Chinese Academy of Sciences, No. 2, Nengyuan Road, Tianhe Area, 510640, Guangzhou, People's Republic of China aut Wang Zhiyuan Key Laboratory of Renewable Energy, Guangzhou Institute of Energy Conversion, Chinese Academy of Sciences, No. 2, Nengyuan Road, Tianhe Area, 510640, Guangzhou, People's Republic of China aut Lv Pengmei Key Laboratory of Renewable Energy, Guangzhou Institute of Energy Conversion, Chinese Academy of Sciences, No. 2, Nengyuan Road, Tianhe Area, 510640, Guangzhou, People's Republic of China aut Yuan Zhenhong Key Laboratory of Renewable Energy, Guangzhou Institute of Energy Conversion, Chinese Academy of Sciences, No. 2, Nengyuan Road, Tianhe Area, 510640, Guangzhou, People's Republic of China aut Journal of Industrial Microbiology & Biotechnology Springer Berlin Heidelberg 42/8(2015-08-01), 1175-1182 1367-5435 42:8<1175 2015 42 10295 https://doi.org/10.1007/s10295-015-1633-6 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s10295-015-1633-6 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- He Dong Key Laboratory of Renewable Energy, Guangzhou Institute of Energy Conversion, Chinese Academy of Sciences, No. 2, Nengyuan Road, Tianhe Area, 510640, Guangzhou, People's Republic of China aut NATIONALLICENCE 700 1- Luo Wen Key Laboratory of Renewable Energy, Guangzhou Institute of Energy Conversion, Chinese Academy of Sciences, No. 2, Nengyuan Road, Tianhe Area, 510640, Guangzhou, People's Republic of China aut NATIONALLICENCE 700 1- Wang Zhiyuan Key Laboratory of Renewable Energy, Guangzhou Institute of Energy Conversion, Chinese Academy of Sciences, No. 2, Nengyuan Road, Tianhe Area, 510640, Guangzhou, People's Republic of China aut NATIONALLICENCE 700 1- Lv Pengmei Key Laboratory of Renewable Energy, Guangzhou Institute of Energy Conversion, Chinese Academy of Sciences, No. 2, Nengyuan Road, Tianhe Area, 510640, Guangzhou, People's Republic of China aut NATIONALLICENCE 700 1- Yuan Zhenhong Key Laboratory of Renewable Energy, Guangzhou Institute of Energy Conversion, Chinese Academy of Sciences, No. 2, Nengyuan Road, Tianhe Area, 510640, Guangzhou, People's Republic of China aut NATIONALLICENCE 773 0- Journal of Industrial Microbiology & Biotechnology Springer Berlin Heidelberg 42/8(2015-08-01), 1175-1182 1367-5435 42:8<1175 2015 42 10295