By: Harsha Gouda

Edited by: Emily Glass, Lisa Pinatti, and Sarah Kearns

 

Vitamins are the essential micronutrients required in tiny amounts for the healthy development of an individual. They play a crucial role by initiating various chemical reactions inside the cells like production of pigment that is responsible for your vision or synthesis of red blood cells that carry oxygen in your blood. Early discovery of the importance of vitamins such as B12 in diet was identified in patients suffering from abnormally large red blood cells. Vitamin B12 is one of the most complex chemical molecules among all the other essential vitamins. Owing to its complexity, this precious and rare vitamin is mainly acquired via diet as humans are unable to synthesize it for ourselves. Low dietary B12 intake or defects in the transport pathway to its target place of utilization inside our body result in heart and nerve related disorders in humans.1 As such, evolution developed a very sophisticated trafficking pathway to transport and deliver B12 from the food we eat to the cells that use it.1

Before it was called B12, this vitamin first was termed as the “liver factor” in 1920, as it was observed that dogs that consumed a diet containing liver helped regenerate their red blood cells 2. The race to identification of the “liver factor” lasted for about three decades until Hodgkin and co-workers isolated and elucidated the chemical composition and structure of vitamin B12 in 1956 using X-ray crystallography.3 They revealed that B12 contains a cobalt metal centre in its structure and it is also the only metal containing B-vitamin in humans.

Activation of B12:

Similar to multiple gadgets you find in the kids toy collection, vitamin B12 is also found in multiple analogues of chemical composition where only a fraction of them are functional. Chemically diverse available B12 could be differentiated into biologically active and inactive forms (Figure). These forms differ in chemical nature of the cobalt metal ion of B12 that is pivotal to its functionality. It is critical to convert the biologically inactive form of B12 to its active form for utilization inside our cells, like the toys that need to be repaired for the fulfilled utilization for the kids. Hence the dietary B12 we consume needs to be processed before it becomes functional. In our diets, we typically ingest B12 in its inactive form as cyano group attached to the cobalt metal centre (Figure). These proteins in our cells termed as “B12 trafficking proteins” , are essential for conversation of B12 into its active form and for its delivery to the target place of utilization. They not only bind and protect B12 from loss due to dilution in our body, they are also critical in its repair and delivery to the target place of utilization. One of B12 trafficking protein in humans, CblC, plays an important role by removing the cyano group attached to the cobalt metal centre from the dietary B12, an essential step in activation of B12. After the detachment of cyano group, B12 can have different chemical groups of varying sizes; for example, small methyl groups or big adenosyl groups that are competent for utilization inside the cell. It is postulated that methyl group is attached to the cobalt centre in B12 by the protein methionine synthase. Methionine synthase in the cytoplasm of our cells uses the methyl group on B12 to perform methyl transfer reactions required for DNA synthesis. However, protein CblB in the mitochondrial compartment of the cells attaches adenosyl group to the cobalt metal in B12.4 Protein mutase in mitochondria uses the adenosyl form of B12 to channel the molecules from breakdown of fatty acids to produce energy. Hence, the B12 is utilized in both cytoplasm and mitochondrial compartment of the cells to perform critical chemical reactions.5 Defects in any of the B12 utilizing or trafficking proteins in humans will result in diseases associated to B12 deficiency.

Utilization of B12:

Once it is active inside the cell, B12 is required for human metabolism via two proteins: methionine synthase and methylmalonyl-CoA mutase. Methionine synthase plays a crucial role in the synthesis of our genetic material, DNA. Methionine synthase is involved in the production of methionine from molecule homocysteine, in doing so it regenerates a pool of tetrahydrofolate molecules essential for DNA synthesis. When it cannot synthesize methionine due to insufficient B12, homocysteine builds up in our body, leading to large and immature red blood cells (homocystinuria).6,7 Methylmalonyl-CoA mutase (referred to as “mutase” here) in the mitochondrial compartment within our cells plays a vital role in the metabolism of fatty acids and amino acids. Break down of fatty acids and amino acids produce ATP molecules, an energy currency inside the cells that can be utilized for various other functions. Importantly, the adenosyl form of B12 allows mutase to produce succinyl-CoA molecule, that will be utilized to produce ATP molecules.8 The adenosyl form of B12 when bound to the mutase breaks a bond that attaches the cobalt metal centre to the adenosyl group (depicted by the green line in figure) to perform its function. Due to leaky nature of the mutase, fraction of B12 used for chemical reaction in mutase results in the formation of inactive form of B12 that can’t be utilized.

Because B12 is a precious commodity in cells, proteins in mitochondria work together to recycle the B12 to an active form after it has been inactivated by the mutase. Two proteins, CblA and CblB, come to the rescue of B12 in mitochondria by repairing and delivering the activated form of B12 back to mutase. Chaperon protein CblA facilitates the movement of inactivated B12 from mutase to CblB. After receiving the inactive B12 from mutase, CblB adds the missing adenosyl group to the cobalt metal centre of B12 using ATP.  Defects in mitochondrial B12 proteins, such as mutase, CblA or CblB result in improper processing of B12 causing elevated levels of methylmalonic acid, a toxic chemical harmful to our body at high concentrations 9. As such, B12 restoration is almost just as important as B12 to our cells. Like humans, many organisms including some bacteria, have their own individual B12 dependent enzymes for survival. Hence, selective inhibition of these bacterial B12 dependent proteins could be leveraged for treating infections. Recently the discovery in immune response leading to inactivation of bacterial mutase function in Mycobacterium tuberculosis has gained a lot of attention to look at Mycobacterium’s mutase as a new drug target to treat tuberculosis infection.10

From the current understanding of the role of vitamin B12 in humans, it is evident that B12 is an important component to be included in everyone’s diet. To increase the amount of B12 in your diet, consuming foods with high B12 amounts like animal meat, dairy products, and eggs are all good options to consider.11 For vegan diet followers, B12 can be obtained as a supplement or from the fortified foods, such as cereals. Although a lot of interesting science is being currently investigated about B12 and B12 dependent proteins, the clinical data has proved B12 as an essential micronutrient to humans.

Figure
Figure 1: Chemical structure of vitamin B12 and its biological active and inactive forms. Representation for utilization of methyl (top-middle) and adenosyl (bottom-middle) form of vitamin B12 in a healthy individual.

 

References:

  1. Watkins, D. & Rosenblatt, D. S. Inborn errors of cobalamin absorption and metabolism. Am. J. Med. Genet. Part C Semin. Med. Genet. 157, 33–44 (2011).
  2. Whipple, G. H., Robscheit, F. S. & Hooper, C. W. BLOOD REGENERATION FOLLOWING SIMPLE ANEMIA. Am. J. Physiol. Content 53, 236–262 (1920).
  3. Hodgkin, D. C. et al. Structure of vitamin B12. Nature 178, 64–66 (1956).
  4. Padovani, D., Labunska, T., Palfey, B. A., Ballou, D. P. & Banerjee, R. Adenosyltransferase tailors and delivers coenzyme B12. Nat. Chem. Biol. 4, 194–196 (2008).
  5. Kim, J., Gherasim, C. & Banerjee, R. Decyanation of vitamin B12 by a trafficking chaperone. Proc. Natl. Acad. Sci. U. S. A. 105, 14551–14554 (2008).
  6. Green, R. et al. Vitamin B12 deficiency. Nat. Rev. Dis. Prim. 3, 17040 (2017).
  7. L N Willoughby M A Pears, B. M., Sharp, A. A. & Shields, J. Megaloblastic Erythropoiesis in Acquired Hemolytic Anemia.
  8. Mancia, F. & Evans, P. R. Conformational changes on substrate binding to methylmalonyl CoA mutase and new insights into the free radical mechanism. Structure 6, 711–720 (1998).
  9. Ledley, F. D., Levy, H. L., Shih, V. E., Benjamin, R. & Mahoney, M. J. Benign Methylmalonic Aciduria. N. Engl. J. Med. 311, 1015–1018 (1984).
  10. Ruetz, M. et al. Itaconyl-CoA forms a stable biradical in methylmalonyl-CoA mutase and derails its activity and repair. Science (80-. ). 366, 589–593 (2019).
  11. Dietary Reference Intakes for Thiamin, Riboflavin, Niacin, Vitamin B6, Folate, Vitamin B12, Pantothenic Acid, Biotin, and Choline. Washington (DC): National Academies Press (US); 1998. 9, Vitamin B12.

 


Harsha Gouda

Harsha completed his integrated bachelor’s and master’s degree at the Indian Institute of science education and research, Pune, India. During his master’s program, he came to the University of Michigan as an international exchange student and he then later decided to join as a Ph.D. student at the department of biological chemistry, UofM, under the guidance of Prof. Ruma Banerjee. Harsha focuses his research studying micro molecular machines, often referred to as enzymes. Besides enjoying his experiments in the lab, he likes to listen to music, playing games (I get competitive sometimes!), swimming in the Huron River and traveling.

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