Cobalamine (B12)

Metabolism of Vitamin B12

1. Cobalamine (B12] Gandham . Rajeev Department of Biochemistry, Akash Institute of Medical Sciences & Research Centre, Devanahalli , Bangalore, Karnataka, India. E-Mail: [email protected]

2. Vitamin B12

3. Cobalamin Extrinsic factor (EF) of castle Antipernicious anemia factor Chemistry: Vitamin B12 is water soluble, heat stable and red in colour It contains 4.35% cobalt by weight Four pyrrole rings co- ordinated with a cobalt atom is called as a Corrin ring Synonyms of Vitamin B12

4. The corrin ring has four pyrrol units, like porphyrin Two of the pyrrole units (A&D) are directly bound to each other where as the other two (B & C) are held by methene bridges The groups namely methyl, acetamide and propionamide are the substituents on the pyrrole rings Cobalt present at the centre of the corrin ring is bonded to the four pyrrole nitrogens

5. Cobalt also holds dimethyl benzimidazole (DMB) containing ribose 5-Phosphate and amino isopropanol A nitrogen atom dimethyl benzimidazole is linked to cobalt The amide group of aminoisopropanol binds with D ring of corrin The cobalt atom also possesses a sixth substituent group located above the plane of the corrin ring

6. A D B C Aminoisopropanol Methyl, Adenosine, acetamide , propionamide

7. Aminoisopropanol dimethyl benzimidazole D B A C

8. All the forms of vitamin B12 are called cobalamins because of the presence of cobalt atom Different forms of cobalamins include Cyanocabalamin :- It refers to the isolated form of vitamin B12 in which cyanide was added to promote crystallization of cobalamin during the isolation process Hydroxycobalamin :- It was usually present in the tissues and in the naturally occurring forms of Vitamin B12

9. Methylcobalamin :- Methyl group replaces adenosyl group Adenosyl cobalamin (Ado-B12):- When taken up by the cells, these groups are removed and deoxy adenosyl cobalamin or Ado-B12 is formed

10. In the food, vitamin B12 is present as a complex with proteins The free form of vitamin B12 is released by cooking, HCL present in gastric juice and proteolysis by pepsin in the stomach Mechanism:- The absorbance of vitamin B12 requires intrinsic factor Intrinsic factor is a glycoprotein secreted by parietal cells of stomach metabolism

11. Vitamin B12 combines with intrinsic factor The vitamin B12 – intrinsic factor complex reaches the ileum where it is absorbed In the ileum, the complex attaches to a specific receptor and is taken up by the mucosal cell. In the mucosal cell, vitamin B12 is released from its complex and reaches the portal circulation

12. In the portal blood, it is transported in combination with transcobalamin II Vitamin B12 is presented to cells where it is taken up by the cells through receptor mediated endocytosis Storage: It is mainly stored in liver, leukocytes and gastric mucosa It is stored as complex with transcobalamin -I&II Transport

13. Protein B12 B12 B12 B12 Protein IF IF IF Mucosal cell B12 Methyl B12 B12 B12 TC 1-B12(90%) TC 1-B12(90%) TCII -B12(10%) TCII -B12(10%) Tissues B12 TCII Methyl B12 Deoxyadenosyl B12 (LIVER) GIT Plasma Tissues

14. About 10 enzymes requiring vitamin B12 have been identified Most of them are found in bacteria ( mutase , ribonucleotide reductase, etc.) There are two reactions in mammals that dependent on vitamin B12 Biochemical functions

15. Methyl cobalamin is essential for the conversion of homocysteine to methionine and formation of FH4 from methyl FH4 The reaction is catalyzed by homocysteine methyl transferase Synthesis of Methionine from homosysteine Homocysteine Homocysteine methyl transferase Methionine methylcobalamin THF N5 methyl THF

16. The circulating methyl FH4 is converted to FH4 FH4 is either used for storage as folylpolyglutamate form or it is utilized for other reactions such as formation of methylene FH4 Methyl folate trap:- In B12 deficiency , impaired conversion of methyl FH4 to FH4 results in accumulation of methyl FH4 & is called as methyl folate trap Methyl folate trap results in decreased availability of FH4 & FH4 derivatives that are Significance of the reaction

17. Needed for purine nucleotide & thymidylate synthesis Thus vitamin B12 deficiency, results in secondary folate deficiency

18. Role of methyl cobalamin & folate trap Methyl FH4 FH4 N5,10 methylene FH4 dUMP dTMP DNA Homocysteine Methionine Methylfolate trap Methyl FH4 Homocysteine methyl transferase FH4 Serine Glycine Thymidylate synthase B12

19. Isomerization of methyl CoA to succinyl CoA:- The degradation of odd chain fatty acids and some amino acids ( valine , leucine etc) and pyrimidines (thymine & Uracil ) produce propionyl CoA, an imp. Compound methylmalonyl CoA The methyl malonyl CoA mutase converts methyl malonyl CoA to succinyl CoA in the presence of Vitamin B12,deoxyadenosyl cobalamin

20. In B12 deficiency , methyl malonyl CoA accumulates and is excreted in urine as methyl malonic acid This condition is called as methylmalonic aciduria , occurs in B12 deficiency. Demyelination :- Myelination of nerves is impaired in B12 deficiency due to accumulation of methylmalonyl CoA Demyelination is due excessive accumulation of methylmalonyl CoA

21. Odd chain FA Propionyl CoA Amino acids (Val, Ile, Thr, Met) Thymine, uracil Methyl malonyl CoA Succinyl CoA Methyl malonyl CoA mutase Methyl malonic acid Excreted in urine 5-Deoxyadenosylcabalamin (of B12) + Methylmalonic aciduria

22. Rich sources of vitamin B12 are meats, egg, milk, sea foods B12 is synthesized by microorganisms Vitamin B12 is not present in Plant sources Dietary sources

23. Adults -1 µg/day Pregnancy & lactation -2 µg/day Causes: Inadequate intake-seen in pure vegetarians and rarely in alcoholism Impaired absorption This is mainly caused by lack of intrinsic factor Lack of intrinsic factor is called as pernicious RDA Deficiency

24. anemia & it is caused by destruction of gastric mucosa Impaired absorption is also seen in small intestinal disorders Impaired storage and transport: Inadequate utilization of vitamin occurs because of liver diseases and abnormalities of transport proteins Increased requirements are seen in hyperthyroidism, infancy & thalassemia Increased excretion occurs in nephrotic syndrome

25. Vitamin B12 deficiency is characterized by Megaloblastic anemia: Feature of megaloblastic anemia include pallor,fatigue , glossitis ( beefy red tongue ) & slight yellow discoloration of the conjunctiva due to increased unconjugated bilirubin Progression of anemia may result in angina & congestive cardiac failure Clinical Features

26. Gastrointestinal dysfunction: GIT epithelial cells are undergoing rapid turnover & dependent on vitamin B12 B12 deficiency results in weight loss & diarrhea Demyelination of nervous tissue Subacute combined degeneration: Damage to nervous system is seen in B12 deficiency

27. There is demyelination affecting cerebral cortex as well as dorsal column & pyramidal tract Symmetrical paresthesia of extremities, alterations of tendon & deep senses & reflexes, loss of position sense, unsteadiness in gait, positive Romberg’s sign & positive Babinski’s sign are seen Achlorhydria : Absence of acid in gastric juice is associated with B12 deficiency

28. Romberg’s sign & positive Babinski’s sign

29. Serum B12: It is quantitated by radio-immunoassay or by ELISA Methyl melonic acid is excreted in urine FIGLU excretion test Peripheral smear: Peripheral blood & bone marrow morphology shows magaloblastic anemia Homocysteinuria : Excretion of homocysteine in urine Assessment of B12 deficiency

30. Schilling test: Radioactive labelled (Cobalt-60) vitamin B12 1 μ g is given orally In gastric atrophy cases, there is no absorption, hence the entire radioactivity is excreted in faeces & radioactivity is not observed in liver If the cause is nutritional deficiency, there will be increased absorption Then radioactivity is noted in the liver region, with very little excretion in feces

31. References Harper’s Biochemistry 25 th Edition. Fundamentals of Clinical Chemistry by Tietz . Text Book of Medical Biochemistry-A R Aroor . Text Book of Biochemistry-DM Vasudevan Text Book of Biochemistry-MN Chatterjea Text Book of Biochemistry- Dr.U.Satyanarana