Engineering Escherichia coli for increased productivity of serine-rich proteins based on proteome profiling.
Variations in proteome profiles of Escherichia coli in response to the overproduction of human leptin, a serine-rich (11.6% of total amino acids) protein, were examined by two-dimensional gel electrophoresis. The levels of heat shock proteins increased, while those of protein elongation factors, 30S ribosomal protein, and some enzymes involved in amino acid biosynthesis decreased, after leptin overproduction. Most notably, the levels of enzymes involved in the biosynthesis of serine family amino acids significantly decreased. Based on this information, we designed a strategy to enhance the leptin productivity by manipulating the cysK gene, encoding cysteine synthase A. By coexpression of the cysK gene, we were able to increase the cell growth rate by approximately twofold. Also, the specific leptin productivity could be increased by fourfold. In addition, we found that cysK coexpression can improve the production of another serine-rich protein, interleukin-12 beta chain, suggesting that this strategy may be useful for the production of other serine-rich proteins as well. The approach taken in this study should be useful in designing a strategy for improving recombinant protein production.
Han
Mee-Jung
MJ
Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering, BioProcess Engineering Research Center, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Republic of Korea.
Jeong
Ki Jun
KJ
Yoo
Jong-Shin
JS
Lee
Sang Yup
SY
D003470:Culture Media; D003547:Cysteine Synthase / Q000235:genetics / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D005818:Genetic Engineering / Q000379:methods*; D006801:Humans; D020738:Leptin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids; D020543:Proteome; D012694:Serine
D003470:Culture Media; D029968:Escherichia coli Proteins; D020738:Leptin; D020543:Proteome; D012694:Serine; D003547:Cysteine Synthase