Expression of codon-optmized phosphoenolpyruvate carboxylase gene from Glaciecola sp. HTCC2999 in Escherichia coli and its application for C4 chemical production.
We examined the expression of the phosphoenolpyruvate carboxylase (PEPC) gene from marine bacteria in Escherichia coli using codon optimization. The codon-optimized PEPC gene was expressed in the E. coli K-12 strain W3110. SDS-PAGE analysis revealed that the codon-optimized PEPC gene was only expressed in E. coli, and measurement of enzyme activity indicated the highest PEPC activity in the E. coli SGJS112 strain that contained the codon-optimized PEPC gene. In fermentation assays, the E. coli SGJS112 produced the highest yield of oxaloacetate using glucose as the source and produced a 20-times increase in the yield of malate compared to the control. We concluded that the codon optimization enabled E. coli to express the PEPC gene derived from the Glaciecola sp. HTCC2999. Also, the expressed protein exhibited an enzymatic activity similar to that of E. coli PEPC and increased the yield of oxaloacetate and malate in an E. coli system.
Department of Chemical and Biomolecular Engineering, Sogang University, Seoul 121-742, South Korea.
Soohyun
S
Park
Seung Pil
SP
Pack
Jinwon
J
Lee
D001483:Base Sequence; D049109:Cell Proliferation; D003062:Codon / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000166:cytology / Q000378:metabolism*; D005285:Fermentation; D015870:Gene Expression; D008293:Malates / Q000378:metabolism*; D008969:Molecular Sequence Data; D062907:Oxaloacetic Acid / Q000378:metabolism*; D010730:Phosphoenolpyruvate Carboxylase / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D020560:Proteobacteria / Q000201:enzymology*; D011995:Recombination, Genetic; D016415:Sequence Alignment
D003062:Codon; D008293:Malates; D062907:Oxaloacetic Acid; C030298:malic acid; D010730:Phosphoenolpyruvate Carboxylase