title: Reduction of Acetate and Lactate Contributed to Enhancement of a Recombinant Protein Production in E. coli BL21.
abstract: Acetate and lactate in growth media are detrimental to the production of Thermus maltogenic amylase (ThMA), a heterologous protein, as well as to the growth of recombinant Escherichia coli. Only 50 mM of acetate or 10 mM of lactate reduced 90% of specific ThMA activity. In this study, mutant E. coli strains blocked in the ackA-pta or ackA-pta and ldh pathways were created, characterized, and assessed for their culture performace in 300 L-scale fermentation. The ackApta and ldh double-mutant strain formed significantly less lactate and acetate, and produced a concomitant increase in the excretion of pyruvate (17.8 mM) under anaerobic conditions. The ackA-pta mutant strain accumulated significant acetate but had an approximately 2-fold increase in the formation of lactate. The ackA-pta and ldh double-mutant strain had superior overall performance in large-scale culture under suboptimal conditions, giving 67% higher cell density and 66% higher ThMA activity compared with those of the control strain. The doublemutant strain also achieved a 179% improvement in volumetric ThMA production.
authors: Department of Chemical Engineering, Konkuk University, Seoul 143-701, Republic of Korea.|Institute of SK-KU Biomaterials, Konkuk University, Seoul 143-701, Republic of Korea.
authors: Tae-Su
authors: TS
authors: Kim
authors: BioNgene Co., Ltd, Seoul 110-521, Republic of Korea.
authors: Hyung-Moo
authors: HM
authors: Jung
authors: BioNgene Co., Ltd, Seoul 110-521, Republic of Korea.
authors: Sang-Yong
authors: SY
authors: Kim
authors: Department of Chemical Engineering, Konkuk University, Seoul 143-701, Republic of Korea.
authors: Liaoyuan
authors: L
authors: Zhang
authors: Department of Chemical Engineering, Konkuk University, Seoul 143-701, Republic of Korea.
authors: Jinglin
authors: J
authors: Li
authors: Department of Chemical Engineering, Konkuk University, Seoul 143-701, Republic of Korea.
authors: Sujan
authors: S
authors: Sigdel
authors: Department of Chemical Engineering, Konkuk University, Seoul 143-701, Republic of Korea.
authors: Ji-Hyun
authors: JH
authors: Park
authors: Institute of SK-KU Biomaterials, Konkuk University, Seoul 143-701, Republic of Korea.
authors: Jung-Rim
authors: JR
authors: Haw
authors: Department of Chemical Engineering, Konkuk University, Seoul 143-701, Republic of Korea.
authors: Jung-Kul
authors: JK
authors: Lee
mesh_terms: D000085:Acetates / Q000378:metabolism*; D003470:Culture Media / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D006026:Glycoside Hydrolases / Q000096:biosynthesis* / Q000235:genetics; D019344:Lactic Acid / Q000378:metabolism*; D060847:Metabolic Engineering; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics; D013824:Thermus / Q000201:enzymology / Q000235:genetics
chemical_list: D000085:Acetates; D003470:Culture Media; D011994:Recombinant Proteins; D019344:Lactic Acid; D006026:Glycoside Hydrolases; C120909:glucan 1,4-alpha-maltohydrolase
keywords: large-scale bioreactor; maltogenic amylase; metabolically engineered; production; recombinant protein