Vitamin B12 (Cobalamin) Bioproduction Engineering Service

Vitamin B12 (Cobalamin) is a complex, essential cofactor used in pharmaceuticals to treat anemia and nerve disorders, and as a supplement in food and animal feed. Production is entirely microbial, but faces challenges: low microbial fermentation yield due to the high energy cost of the 30-step biosynthetic pathway and tight self-regulatory mechanisms in the host (Propionibacterium). Furthermore, B12 is an intracellular product, leading to difficult separation and purification from the complex cell matrix. This necessitates enhanced production and simplified recovery.

CD Biosynsis offers a synthetic biology service focused on maximizing Vitamin B12 yield and simplifying recovery. Our core strategy involves modification of metabolic regulation in Propionibacterium (the industry standard producer), specifically disrupting the feedback inhibition exerted by high Cobalamin levels and enhancing precursor supply (e.g., DMB). This is coupled with the optimization of Cobalamin synthesis pathway by overexpressing rate-limiting enzymes and, crucially, engineering the host for extracellular secretion of B12. Enhanced secretion bypasses the need for cell lysis and greatly simplifies downstream purification. This integrated approach aims to deliver a high-yield, pure, and economically viable naturally produced Vitamin B12 product.

Pain Points

Maximizing the efficiency of Vitamin B12 bioproduction faces these critical challenges:

• Low Microbial Fermentation Yield: The B12 pathway is extremely long (approx. 30 genes) and energetically expensive. Expression is tightly regulated by a Cobalamin riboswitch that shuts down transcription when B12 levels are high, limiting yield.
• Difficult Separation and Purification: B12 is primarily an intracellular product in Propionibacterium. Recovery requires intensive cell lysis (often heat/acid treatment) followed by complex chromatographic steps, leading to high cost and potential degradation.
• DMB Precursor Supply: The required precursor, Dimethylbenzimidazole (DMB) , is often the rate-limiting component, requiring co-fermentation or external addition.
• Product Form Heterogeneity: Fermentation produces hydroxocobalamin, which must be converted to cyanocobalamin (the stable commercial form), adding an extra post-fermentation step.

A successful solution must break the regulatory feedback loop, boost pathway flux, and enable efficient product secretion.

Solutions

CD Biosynsis utilizes advanced metabolic engineering to optimize Vitamin B12 production in Propionibacterium and related hosts:

Modification of Metabolic Regulation in Propionibacterium

           

We disrupt the B12 riboswitch to abolish feedback inhibition, ensuring continuous high-level expression of the Cobalamin synthesis gene cluster.

Optimization of Cobalamin Synthesis Pathway

We perform multi-gene cluster overexpression (using strong constitutive promoters) focusing on rate-limiting steps, particularly the final adenosylation and salvage pathways.

Extracellular Secretion Engineering

We introduce or engineer B12-specific membrane transporters or modify membrane structure to enable the efficient export of the product into the fermentation broth.

Precursor Pathway Balancing

We modify the host’s purine metabolism to boost the de novo synthesis of DMB (the lower ligand) from simple carbon sources, removing the need for external DMB supplementation.

This systematic approach is focused on rebuilding the host’s regulatory and synthetic machinery to achieve unprecedented yield and simplified downstream processing.

Advantages

Our Vitamin B12 engineering service is dedicated to pursuing the following production goals:

Significantly Increased Titer

Riboswitch disruption and pathway boosting overcome natural yield limitations, leading to high industrial production titers .

Reduced Purification Costs

B12 secretion into the broth avoids costly cell lysis steps , dramatically simplifying downstream purification.

Simplified Feedstock Needs

DMB precursor pathway engineering eliminates the need for expensive external supplementation , lowering raw material costs. [Image of Cost Reduction Icon]

Robust Fermentation Host

Utilizing Propionibacterium or related GRAS strains provides a well-established, safe industrial platform .

High Bioavailability

The biosynthetic product is the active, naturally occurring form (hydroxocobalamin) before final conversion.

We provide a specialized platform aimed at maximizing the yield and minimizing the cost of Vitamin B12 biomanufacturing.

Process

Our Vitamin B12 strain engineering service follows a rigorous, multi-stage research workflow:

• Riboswitch Engineering: Modify or delete the regulatory riboswitch structure in the leader sequence of the Cobalamin gene cluster to achieve constitutive expression.
• Pathway Overexpression: Use strong, stable promoters to overexpress Cobalamin synthesis genes (the Cbi and Cob clusters) to boost flux.
• DMB Precursor Supply: Engineer the purine synthesis pathway to enhance the flow of nucleotide precursors toward DMB synthesis.
• Secretion System Construction: Introduce or activate a B12-specific ABC transporter system to efficiently pump the product into the medium.
• Fermentation Performance Validation: Test the final engineered strain in optimized anaerobic fermentation to assess B12 titer (intracellular and extracellular), yield, and stability .
• Result Report Output: Compile a detailed Experimental Report including gene modification data, flux analysis, and fermentation metrics (yield, titer, and purity) , supporting commercial scale-up.

Technical communication is maintained throughout the process, focusing on timely feedback regarding yield and product secretion efficiency.

Explore the potential for a high-yield, cost-effective Vitamin B12 supply. CD Biosynsis provides customized strain engineering solutions:

• Detailed Titer and Secretion Analysis Report , demonstrating the degree of yield enhancement and extracellular accumulation.
• Consultation on post-fermentation conversion (e.g., hydroxocobalamin to cyanocobalamin) and primary capture steps.
• Experimental reports include complete raw data on volumetric productivity (mg/L/h) and product purity , essential for pharmaceutical and food grade applications.