Identification of riboflavin: revealing different metabolic characteristics between Escherichia coli BL21(DE3) and MG1655.
There are many physiological differences between Escherichia coli B and K-12 strains, owing to their different origins. Deeper insight into the metabolic and regulative mechanisms of these strains will inform improved usage of these industrial workhorses. In the present study, we observed that BL21 fermentation broth gradually turned yellow during cultivation. By spectral analysis and liquid chromatography-mass spectrometry identification, we confirmed for the first time that the yellow substance accumulated in the fermentation broth is riboflavin. Comparing the enzyme sequences involved in riboflavin metabolism between BL21 and MG1655, we identified a site mutation on the 115 residue of bifunctional riboflavin kinase/FMN adenylyltransferase (RibF) in BL21. This His115Leu mutation was found to reduce enzyme activity to 55% of that of MG1655, which is probably one reason for riboflavin accumulation in BL21. Quantitative PCR analysis showed that genes of the entire branch of the riboflavin and FAD biosynthesis pathways in BL21 were up-regulated. Several physiological and metabolic characteristics of BL21 and MG1655 were found to be different, and may also be related to the riboflavin accumulation.
National Glycoengineering Research Center, State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, P. R. China.
Xinran
X
Wang
National Glycoengineering Research Center, State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, P. R. China qiqi20011983@gmail.com.
Qian
Q
Wang
National Glycoengineering Research Center, State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, P. R. China.
Qingsheng
Q
Qi
D004926:Escherichia coli / Q000145:classification / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D048168:Escherichia coli K12 / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D005285:Fermentation; D005182:Flavin-Adenine Dinucleotide / Q000235:genetics / Q000378:metabolism; D008401:Gas Chromatography-Mass Spectrometry; D009154:Mutation; D009713:Nucleotidyltransferases / Q000235:genetics; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D012256:Riboflavin / Q000302:isolation & purification / Q000378:metabolism*
D005182:Flavin-Adenine Dinucleotide; D017853:Phosphotransferases (Alcohol Group Acceptor); C022532:riboflavin kinase; D009713:Nucleotidyltransferases; C020143:FMN adenylyltransferase; D012256:Riboflavin
BL21(DE3); Escherichia coli; MG1655; metabolic difference; riboflavin