Development and application of an arabinose-inducible expression system by facilitating inducer uptake in Corynebacterium glutamicum.
Corynebacterium glutamicum is currently used for the industrial production of a variety of biological materials. Many available inducible expression systems in this species use lac-derived promoters from Escherichia coli that exhibit much lower levels of inducible expression and leaky basal expression. We developed an arabinose-inducible expression system that contains the L-arabinose regulator AraC, the P(BAD) promoter from the araBAD operon, and the L-arabinose transporter AraE, all of which are derived from E. coli. The level of inducible P(BAD)-based expression could be modulated over a wide concentration range from 0.001 to 0.4% L-arabinose. This system tightly controlled the expression of the uracil phosphoribosyltransferase without leaky expression. When the gene encoding green fluorescent protein (GFP) was under the control of P(BAD) promoter, flow cytometry analysis showed that GFP was expressed in a highly homogeneous profile throughout the cell population. In contrast to the case in E. coli, P(BAD) induction was not significantly affected in the presence of different carbon sources in C. glutamicum, which makes it useful in fermentation applications. We used this system to regulate the expression of the odhI gene from C. glutamicum, which encodes an inhibitor of α-oxoglutarate dehydrogenase, resulting in high levels of glutamate production (up to 13.7 mM) under biotin nonlimiting conditions. This system provides an efficient tool available for molecular biology and metabolic engineering of C. glutamicum.
Department of Industrial Microbiology and Biotechnology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China.
Yun
Y
Zhang
Xiuling
X
Shang
Shujuan
S
Lai
Guoqiang
G
Zhang
Yong
Y
Liang
Tingyi
T
Wen
D001089:Arabinose / Q000378:metabolism*; D002244:Carbon / Q000378:metabolism; D048230:Corynebacterium glutamicum / Q000235:genetics* / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics; D005434:Flow Cytometry; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects*; D017930:Genes, Reporter; D005822:Genetic Vectors; D005827:Genetics, Microbial / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000235:genetics; D008967:Molecular Biology / Q000379:methods; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000235:genetics
D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D014157:Transcription Factors; D049452:Green Fluorescent Proteins; D002244:Carbon; D001089:Arabinose