Enhancing Production of Bio-Isoprene Using Hybrid MVA Pathway and Isoprene Synthase in E. coli

The depleting petroleum reserve, increasingly severe energy crisis, and global climate change are reigniting enthusiasm for seeking sustainable technologies to replace petroleum as a source of fuel and chemicals. In this paper, the efficiency of the MVA pathway on isoprene production has been improved as follows: firstly, in order to increase MVA production, the source of the “upper pathway” which contains HMG-CoA synthase, acetyl-CoA acetyltransferase and HMG-CoA reductase to covert acetyl-CoA into MVA has been changed from Saccharomyces cerevisiae to Enterococcus faecalis; secondly, to further enhance the production of MVA and isoprene, a alanine 110 of the mvaS gene has been mutated to a glycine. The final genetic strain YJM25 containing the optimized MVA pathway and isoprene synthase from Populus alba can accumulate isoprene up to 6.3 g/L after 40 h of fed-batch cultivation.

[1]  J. Keasling,et al.  Engineering a mevalonate pathway in Escherichia coli for production of terpenoids , 2003, Nature Biotechnology.

[2]  M. Schwarz Terpen-Biosynthese in Ginkgo biloba , 1994 .

[3]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[4]  Jay D Keasling,et al.  Microbial sensors for small molecules: development of a mevalonate biosensor. , 2007, Metabolic engineering.

[5]  A. Bacher,et al.  Biosynthesis of isoprenoids via the non-mevalonate pathway , 2004, Cellular and Molecular Life Sciences CMLS.

[6]  Autumn L. Sutherlin,et al.  X-ray crystal structures of HMG-CoA synthase from Enterococcus faecalis and a complex with its second substrate/inhibitor acetoacetyl-CoA. , 2005, Biochemistry.

[7]  M. Rodríguez-Concepcíon,et al.  Elucidation of the Methylerythritol Phosphate Pathway for Isoprenoid Biosynthesis in Bacteria and Plastids. A Metabolic Milestone Achieved through Genomics1 , 2002, Plant Physiology.

[8]  In Vitro Assembly of Multiple DNA Fragments Using Successive Hybridization , 2012, PloS one.

[9]  M. Rodríguez-Concepcíon,et al.  The MEP pathway: a new target for the development of herbicides, antibiotics and antimalarial drugs. , 2004, Current pharmaceutical design.

[10]  E. Cox,et al.  Site-specific chromosomal integration of large synthetic constructs , 2010, Nucleic acids research.

[11]  Jay D Keasling,et al.  Balancing a heterologous mevalonate pathway for improved isoprenoid production in Escherichia coli. , 2007, Metabolic engineering.

[12]  Autumn L. Sutherlin,et al.  A structural limitation on enzyme activity: the case of HMG-CoA synthase. , 2006, Biochemistry.

[13]  J. Sacchettini,et al.  Creating Isoprenoid Diversity , 1997, Science.

[14]  C. N. Hewitt,et al.  Exposure to isoprene promotes flowering in plants , 1995 .

[15]  Mattijs K. Julsing,et al.  Functional analysis of genes involved in the biosynthesis of isoprene in Bacillus subtilis , 2006 .

[16]  A. Margaritis,et al.  Airlift-driven fibrous-bed bioreactor for continuous production of glucoamylase using immobilized recombinant yeast cells. , 2009, Journal of biotechnology.

[17]  E. Eroğlu,et al.  Extracellular terpenoid hydrocarbon extraction and quantitation from the green microalgae Botryococcus braunii var. Showa. , 2010, Bioresource technology.

[18]  J. Sohng,et al.  Improved Squalene Production via Modulation of the Methylerythritol 4-Phosphate Pathway and Heterologous Expression of Genes from Streptomyces peucetius ATCC 27952 in Escherichia coli , 2009, Applied and Environmental Microbiology.

[19]  J. Peñuelas,et al.  Linking isoprene with plant thermotolerance, antioxidants and monoterpene emissions , 2005 .

[20]  James M Clomburg,et al.  Biofuel production in Escherichia coli: the role of metabolic engineering and synthetic biology , 2010, Applied Microbiology and Biotechnology.

[21]  Junfeng Xue,et al.  Enhancing Isoprene Production by Genetic Modification of the 1-Deoxy-d-Xylulose-5-Phosphate Pathway in Bacillus subtilis , 2011, Applied and Environmental Microbiology.

[22]  Jay D Keasling,et al.  Optimization of the mevalonate-based isoprenoid biosynthetic pathway in Escherichia coli for production of the anti-malarial drug precursor amorpha-4,11-diene. , 2009, Metabolic engineering.

[23]  H. Sahm,et al.  Isoprenoid biosynthesis in bacteria: a novel pathway for the early steps leading to isopentenyl diphosphate. , 1993, The Biochemical journal.

[24]  A. Steinbüchel Production of rubber-like polymers by microorganisms. , 2003, Current opinion in microbiology.

[25]  A. Melis,et al.  Engineering a platform for photosynthetic isoprene production in cyanobacteria, using Synechocystis as the model organism. , 2010, Metabolic engineering.

[26]  Jay D Keasling,et al.  High‐level production of amorpha‐4,11‐diene in a two‐phase partitioning bioreactor of metabolically engineered Escherichia coli , 2006, Biotechnology and bioengineering.

[27]  Zhinan Xu,et al.  Enhanced plasmid stability and production of hEGF by immobilized recombinant E. coli JM101 , 2006 .

[28]  Antia Rodriguez-Villalon,et al.  Carotenoid accumulation in bacteria with enhanced supply of isoprenoid precursors by upregulation of exogenous or endogenous pathways. , 2008, Journal of biotechnology.

[29]  S. Sauret-Güeto,et al.  Escherichia coli engineered to synthesize isopentenyl diphosphate and dimethylallyl diphosphate from mevalonate: a novel system for the genetic analysis of the 2-C-methyl-d-erythritol 4-phosphate pathway for isoprenoid biosynthesis. , 2001, The Biochemical journal.

[30]  S. Hashimoto,et al.  Production of mevalonate by a metabolically-engineered Escherichia coli , 2004, Biotechnology Letters.

[31]  F. Loreto,et al.  Isoprene produced by leaves protects the photosynthetic apparatus against ozone damage, quenches ozone products, and reduces lipid peroxidation of cellular membranes. , 2001, Plant physiology.

[32]  Shao-Yi Hou,et al.  Multiple-copy-gene integration on chromosome of Escherichia coli for beta-galactosidase production , 2008 .

[33]  Mo Xian,et al.  Biosynthesis of isoprene in Escherichia coli via methylerythritol phosphate (MEP) pathway , 2011, Applied Microbiology and Biotechnology.

[34]  Jay D Keasling,et al.  Combinatorial expression of bacterial whole mevalonate pathway for the production of beta-carotene in E. coli. , 2009, Journal of biotechnology.

[35]  Myoung-Dong Kim,et al.  Amplification of 1-deoxy-d-xyluose 5-phosphate (DXP) synthase level increases coenzyme Q10 production in recombinant Escherichia coli , 2006, Applied Microbiology and Biotechnology.

[36]  M. Rohmer,et al.  Isoprenoid biosynthesis in Escherichia coli via the methylerythritol phosphate pathway: enzymatic conversion of methylerythritol cyclodiphosphate into a phosphorylated derivative of (E)-2-methylbut-2-ene-1,4-diol , 2002 .