Methionine Bioproduction Engineering Service

Methionine (Met) is an essential sulfur-containing amino acid widely used as a critical supplement in the animal feed industry and in human nutrition. While chemical synthesis (producing DL-Methionine) is dominant, it is highly problematic, resulting in high pollution in chemical synthesis due to the use of toxic chemicals and waste generation. Biosynthesis offers an environmentally superior route but faces challenges from complex biosynthetic pathways involving multi-step conversions, stringent regulation, and reliance on expensive cofactors like S-Adenosylmethionine (SAM).

CD Biosynsis offers a synthetic biology service focused on engineering Escherichia coli for high-titer L-Methionine production. Our core strategy involves modification of methionine synthesis pathway in Escherichia coli . The Methionine pathway branches off the Aspartate pathway. Key enzymes involved include Homoserine O-succinyltransferase (MetA), which is the first committed step and subject to feedback inhibition, and Methionine synthase (MetH or MetE). We engineer the pathway to overcome these regulatory hurdles, primarily through the overexpression of key enzymes . We amplify the expression of pathway enzymes, focusing specifically on MetA and MetH (or MetE), using strong promoters and high-copy plasmids. Crucially, we use site-directed mutagenesis on MetA to eliminate Methionine feedback inhibition, thereby opening the metabolic faucet. Furthermore, we eliminate competing pathways (e.g., Lysine, Threonine) that divert the common Aspartate precursor, ensuring maximum carbon flux is directed towards L-Methionine. This integrated approach achieves a clean, high-yield biosynthetic process that can compete economically with conventional chemical methods.

Pain Points

Achieving sustainable and cost-effective L-Methionine production faces these key challenges:

• High Pollution in Chemical Synthesis: Chemical production relies on Acrolein and Methylmercaptan, generating toxic and persistent chemical waste that is costly to treat and environmentally damaging.
• Complex Biosynthetic Pathways: The Methionine pathway is highly regulated by feedback inhibition (at MetA) and complex transcriptional regulation (MetJ repressor), making native production low-yield.
• Pathway Leakage: The Aspartate common precursor is diverted to synthesize Threonine, Lysine, and Isoleucine, leading to significant carbon loss from the Methionine pathway.
• Cofactor Dependence: The critical Methionine synthase (MetH) requires the costly and unstable cofactor Vitamin B12 , adding complexity and cost to the fermentation process.

A successful solution must remove regulatory bottlenecks and competing pathways while simplifying cofactor dependence.

Solutions

CD Biosynsis utilizes advanced metabolic and enzyme engineering to optimize Methionine production in E. coli:

Modification of Methionine Synthesis Pathway in E. coli

           

We mutate MetA to eliminate feedback inhibition by Methionine and SAM, and delete the MetJ repressor to ensure constitutive pathway expression.

Overexpression of Key Enzymes

We amplify the expression of the entire pathway, focusing on the rate-limiting steps: MetA^mut, MetB (Cystathionine gamma-synthase), and the B12-independent MetE (Methionine synthase).

Competing Pathway Blockade

We delete Threonine and Lysine pathway genes (e.g., ThrB or LysC) that compete with Met for the common Aspartate precursor.

Cofactor Simplification (MetE Utilization)

We utilize and overexpress the B12-independent MetE enzyme to circumvent the need for the expensive Vitamin B12 cofactor required by MetH.