Background

Vitamin B₁₂ is involved in metabolic functions in all organisms, and is therefore an essential nutrient. However, it can only be synthesized by some bacteria and archaea, thus, animals have to obtain it through their diet. Thereby, only animal products like meat and dairy contain B₁₂ in general. This B₁₂ was synthesized by microorganisms colonizing the gastrointestinal tract of those animals and accumulated in the animals’ tissues. Vitamin B₁₂ is one of the most expensive biochemicals in the world, and its synthesis is extraordinarily complex. Since the chemical production of Vitamin B₁₂ requires 70 synthesis steps, it is far too technically challenging and expensive. Therefore, its production is facilitated by genetically engineered microorganisms. These are able to produce Vitamin B₁₂ in industrial amounts and achieve a high product quality. However, the produced Vitamin B₁₂ is harvested by cell lysis, which prevents a continuous production. The efficiency of production could be increased by exporting Vitamin B₁₂ outside the cells. To date, a natural cellular Vitamin B₁₂ exporter is unknown.

Relevance

An adult needs approximately 3.0 µg Vitamin B₁₂ per day which is essential for certain functions: it is involved in processes concerning synthesis of DNA, hormones and neurotransmitters and is involved in the formation of the nervous system and blood. Hence, a B₁₂ deficiency can cause diverse diseases like cancer, dementia, depression, pernicious anemia and polyneuropathy.

Nowadays, many people do not eat meat or animals products at all. Therefore, these people have a high risk to suffer from B₁₂ deficiency if their diet does not provide enough B₁₂.

Vitamin B₁₂ is also used in the industry as it is needed in the biotechnical production of various organic substances. Furthermore, it is added to diverse daily products like toothpaste, fruit gum, non-diary milk or cleaning solution for contact lenses.

Our Aim

We intend to design, construct and introduce a synthetic Vitamin B₁₂ exporter (“Synporter”) into a production organism. Thereby, we aim for facilitated and higher yields in the industrial Vitamin B₁₂ production without requiring cell lysis. The B₁₂ Synporter consists of a signal for a Twin Arginine Transporter (Tat) mediated export that is linked to a B₁₂-binding domain. This construct will be expressed in S. typhimurium TA100, R. planticola and S. blattae. We will not only test these different production organisms but also different B₁₂-binding domains.