1. Production of vitamin B12 in escheria coli
Ida Ruuth

2. Production of vitamin B12
Vitamin B12 is widely used in medical industry and food industry
Need for large scale industrial production
Usually produced by Pseudomonas denitrificans, Propionibacterium shermanii, or Sinorhizobium meliloti
Long fermentation cycles, complex and expensive media requirements and lack of suitable genetic systems for strain engineering

3. E. coli Fast growing, no expensive or complex media requirements
Well-known
Easy to modify genetically
Has ability to produce ALA via C5 pathway
Precursor for vitamin B12
 E. coli lacks about 20 genes required to synthesize the central ring structure of vitamin B12 
 Has no ability to synthesize vitamin B12 compounds de novo

4. Pathway of adenosylcobalamin
Complex pathway
Fusion enzyme CysG is shared between cobalamin and siroheme synthesis.
Cobalamin riboswitches regulate cobalamin biosynthesis and transportation at the transcriptional or translational level
 btuB gene of E. coli 

5. Production of vitamin B12 in E.coli
More effective ALA supply
Expression of HemA from R. palustris
Building biosynthetic pathway by using pathway tuning and functional components
Genes involved in B12 vitamin pathway into E. coli from S. typhimurium
Optimization of pathways
Regulation
btuB gene

6. Challenges and Problems
How introduced genes function in E. coli
Complex pathway and complex structure of vitamin B12
Balance of expressed genes and enzymes
Challenges in expressing a great amount of recombinant genes
More than 20 genes are involved
Balanced supply of precursors

7. Previous studies
Expression of the operon pduBAF-pocR-cbiABCDETFGHJKLMNQOP- cobUST from S. typhimurium into E. coli strain
 de novo cobalamin biosynthesis
 By Ko et al. twenty-two were cloned via restriction and ligation into three compatible plasmids under the control of a T7 promoter and introduced into E. coli strain.
 Vitamin B12 was produced de novo under both anaerobic and aerobic conditions

8. References
Dmitry A. Rodionov et al Comparative Genomics of the Vitamin B12 Metabolism and Regulation in Prokaryotes. The Journal of Biological Chemistry. 278, DOI: /jbc.M Huan Fang et al Microbial production of vitamin B12: a review and future perspectives. Microbial Cell Factories. DOI: /s y Zhang, L., Chen, J., Chen, N. et al Cloning of two 5-aminolevulinic acid synthase isozymes HemA and HemO from Rhodopseudomonas palustris with favorable characteristics for 5-aminolevulinic acid production. Biotechnol Lett. 35: 763. doi: /s