Vitamin B12 Deficiency: What are the consequences?

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Methylation, vitamin B12, the Folate Cycle and Methylation Mutations

MTHFR

 Normal cycling of folate results in the formation of an irreversible reaction in which 5, 10 methylenetetrahydrofolate is converted to 5-methyltetrahydrydrofolate (5-MTHF) by the enzyme 5,10 methyltetrahydrofolate reductase (MTHFR).

5,10-THF + MTHFR => 5-MTHF

The MTHFR uses FAD (derived from riboflavin, vitamin B2) as an essential cofactor. In addition, the reduction that occurs requires input from NADH + H+ (derived from nicotinamide, vitamin B3). In the absence of these two vitamins (B2/B3) the enzyme is not functional. Individuals who have hypothyroidism and who cannot convert riboflavin to FAD have functionally deficient MTHFR. Normally MTHFR activity is regulated by the levels of SAM, which exhibits feed-back control on MTHFR.

Mutations in the MTHFR gene are associated with elevated homocysteine, which is in turn associated with an increased risk in cardiovascular disease, as well as regional brain atrophy..

Common mutations are C677T and A1298C. In individuals with these mutations, the pathway from the folate cycle into the methylation cycle is greatly restricted, and supplementation with folate has been shown to be ineffective in totally lowering homocysteine levels. Thus individuals with the MTHFR 677TT or 1298CC genome must instead use 5-MTHF. Persons with MTHFR 667TT and 1298CC have been shown to have an increased risk of breast, liver and prostate cancer and an increased risk of MS, PD, cleft palate, Down's syndrome, schizophrenia and infertility. Mutations in MTHFR are often seen in people with CFS and children with ASD

 

MTR

In order for the methyl group "acquired" during the folate cycling to enter the methylation cycle, vitamin B12 (Co(I)-Cbl) associated with methionine synthase accepts the methyl group from 5MTHF which then forms both methyl-cobalamin (MeCbl) and tetrahydrofolate (THF). THF can then enter the folate cycle. Methionine synthase (MTR)  then transfers the methyl group from MeCbl to homocysteine, which is converted to methionine, and Co(I)Cbl is reformed.

5-THF + Co(I)Cbl-MTR => MTHF + MeCbl-MTR

Hcy + MeCbl-MTR => Met + Co(I)Cbl-MTR

The most common mutation in the MTR gene is MTR A2756G. The MTR 2756GG mutation is associated with an increased risk of breast cancer, reduced fertility and hyperhomocysteinemia. Many children with ASD have a reduced activity of this enzyme.

 

MTRR

During the processing of MeCbl the cobalt atom is converted from Co(III) to Co(I). In persons with mutations in the MTHFR gene, the enzyme has reduced activity and so the amount of 5MTHF that is produced from the folate cycle is reduced, and so the 5MTHF + Co(I)Cbl <=> THF + MeCbl reaction cannot operate efficiently. As a result new/incoming supplies of MeCbl must be used for processing homocysteine, leading to a more rapid reduction in vitamin B12 levels. (The extent of reduction is dependent upon how many and which types of mutations occur in the MTHFR and other methylation associated genes). Every methyl group used in synthesis must be supplied by MeCbl. This leads to rapid depletion of VB12 stores. An alternative solution is to supply 5MTHF and Co(I)+Cbl, but as this also is not recycled a new molecule of 5MTHF must always be supplied or the newly synthesized Co(I)-Cbl also will run out. Thus in these individuals they need a constant supply of both MeCbl AND 5MTHF.

Methionine synthase reductase enzyme (MTRR) uses FMN and FAD (derived from riboflavin, vitamin B2) as an essential cofactor. In addition, the reduction that occurs requires input from NADPH + H+ (derived from nicotinamide, vitamin B3). In the absence of these two vitamins (B2/B3) the enzyme is not functional. Individuals who have hypothyroidism and who cannot convert riboflavin to FMN and FAD have functionally deficient MTRR.

During the recycling of Co-MTR occasionally the Co(I) is oxidized to Co(II), which is inactive in the MTR enzyme. MTRR is involved in firstly reducing the Co(ii) to Co(I) and then uses the Methyl group on AdoMet to convert the Co(I) to MeCo(III)-MTR.

Co(II)Cbl-MTR + MTRR + NADH + AdoMet => Co(III)MeCbl-MTRR + NAD+ AdoHcy

The most common mutation in MTRR is MTRR A66G. the MTRR 66GG genotype is associated with increased risk of breast cancer, colorectal carcinoma, short term memory problems, elevated homocysteine, PS, coronary heart disease, NTD, Down syndrome, cleft palate, decreased DNA methylation, and Crohn Disease. Mutations in MTRR are often seen in people with CFS and children with ASD.

The enzymes, MTRR and decyanase are involved in removal of the CN group from CN-Cbl. Lack of vitamins B2, B3 or the presence of the MTRR 66GG protein, means that CNCbl cannot be converted to MeCbl and AdoCbl. 

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CBS

The cystathione B -synthase enzyme (CBS) is involved in moving the sulfur atom from homocysteine into the transulfuration pathway leading to the production of glutathione. In this process homocysteine is converted to cystathione by conjugation to serine. The enzyme uses pyridoxal-5-phosphate as a co-factor (derived from vitamin B6).

Homocysteine + Serine + CBS + P5P => Cystathione +H2O

The CBS enzyme is an important first step in the production of cysteine from dietary methionine, and the formation of glutathione (GSH). The enzyme is controlled by the intracellular concentration of SAM, which binds to the enzyme and increases its activity. The enzyme is also controlled by a heme molecule, which can be oxidized from Fe2+ to Fe3+ particularly under acidic conditions within the cell. The enzyme is inhibited by CO or NO, due to binding to the heme group.

The most common mutations in CBS are CBS A13637G, C669T, A360G, N212G, and C19150T. The V168M mutation results in a 7-fold decrease in bound P5P and 13-fold decrease in activity. The S466L mutation is constitutively activated and is not further activated by SAM. The I278T, G307S, G305R and A114V mutations all respond to increased pyridoxine. Mutations in CBS gene are common in CFS and children with ASD. They are also associated with homocysteinemia, mental retardation (and Down's syndrome), lens dislocation, skeletal abnormalities and vascular disease.

BHMT

Betaine hydroxymethyl transferase provides another pathway for the synthesis of methyl groups for synthesis of SAM.

Homocysteine + Betaine + BHMT + P5P => Methionine +dimethylglycine

The enzyme is found mainly in the liver and kidneys. The activity of BHMT is regulated by SAM concentrations, and hence is inhibited by high concentrations of SAM.

 

Histamine Break-down

The two main enzymes involved in Histamine break-down in the body are HNMT and DAO

HNMT

Histamine-N-Methyltransferase  is the main enzyme involved in break-down of histamine in most tissues. The enzyme requires SAM. It is also the main enzyme for histamine break-down in the Central Nervous System.

Histamine + HNMT +SAM => N-Methyl-histamine + SAH

Mutations in the NMT gene are associated with  myasthenia gravis, Parkinson's disease, multiple sclerosis, and the incidence of migraine.

DAO

Diamino-oxidase is the enzyme involved in Histamine break-down. Mutations in the gene involve the binding site for FAD. Persons with DAO mutations often have a high sensitivity to histamine in food, and can be "hypersensitive".

Histamine + DAO + FAD O2 + H2O => Imidazole acetaldehyde +FADH2 + H2O2 + NH3

 It is mainly found in the kidney, gut, lung and brain.Intestinal DAO is responsible for break-down of histamine in food. Mutations in DAO are associated with the severity of ulcerative colitis. Common mutations showing reduced DAO activity include rs2052129, rs2268999, rs10156191 and rs1049742

MAO

Monoamino-oxidase is the enzyme involved in the break-down of dopamine, nor-epinephrine, epinephrine and serotonin. Mutations in the X-linked gene involve the binding site for FAD. MAO mutations have been associated with ADHD, aggressive behaviour, borderline mental retardation, autism, sleep disorders, and mood swings. This is presumably due to a build up of dopamine, serotonin, epinephrine and 5-HT..

DOPA + MAO + FAD => DHPA+FADH2

Other symptoms of reduced MAO activity include hypotension, nausea, dizziness, dry mouth, sexual dysfunction, headaches, drowsiness, difficulty sleeping, RLS, muscle aches, and pins and needles.

COMT

Catechol-O-methyltransferase is the enzyme involved in the metabolism of DOPA, catecholamines (dopamine, nor-epinephrine and epinephrine), catecholestrogens and ascobic acid.,  Low COMT activity is associated with OCD, aggressive highly anti-social schizophrenia, late onset alcoholism, and bipolar manic depressive disorder.

GAD

Glutamate decarboxylase catalyses the decarboxylation of glutamate to GABA using PLP as a co-factor.  Mutations in GAD have been associated with cerebral palsy, panic disorder in females, heroin addiction, higher BMI in late childhood and adolescence in girls, and schizophrenia.