Sunday, April 29, 2007

Construction of L-threonine producing strain based on in silico simulation and comparative transcriptome analysis

Kwang Ho Lee, Jin Hwan Park, Tae Yong Kim, Hyohak Song, and Sang Yup Lee. Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering, Korea Advanced Institute of Science and Technology, 373-1 Guseong-dong, Yuseong-gu, Daejeon, South Korea

Systems metabolic engineering approaches were applied to elucidate crucial factors affecting the biosynthesis of L-threonine in Escherichia coli. Based on the analysis, various engineered strains were developed by introducing targeted genetic modifications into prototrophic E. coli W3110. The first L-threonine-producing strain was constructed by releasing regulatory mechanisms, such as feedback inhibition and attenuation. This strain carries point mutations in the ilvA gene. The lysA, metA and tdh genes were knocked-out to increase the availability of L-threonine precursors. This model strain was employed for the production of L-threonine by fed-batch fermentation.
Next, we carried out comparative transcriptome analysis to identify the effect of L-threonine overproduction on the cellular physiology. Also, in silico simulation studies were carried out to identify the knock-out and amplification gene targets for enhancing L-threonine production. Based on the results obtained from the comparative transcriptome analysis and in silico simulation studies, further metabolic engineering of the L-threonine producing model strain was conducted. As a result, we we were able to identify crucial factors that otherwise would have been difficult, if not impossible, to be identified.
[This work was supported by a grant from the Korean Ministry of Science and Technology (Korean Systems Biology Research Grant, M10309020000-03B5002-00000). Further support by the LG Chem Chair Professorship is appreciated.]