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methyleneH4folate
formylH4folate
methylH4folate
Metabolic Pathways
Folates: folate → dihydrofolate → tetrahydrofolate (THF) ↔ methylene-THF → methyl-THF
• Methylation regulates gene expression and maintains the
differentiated state.
•The immediate methyl donor in most reactions is Sadenosyl methionine, or SAM. The waste product is Sadenosyl homocysteine. This is converted back to
methionine through a series of reactions involving vitamin
B12 and tetrahydrofolate. Once the homocysteine has
been re-methylated, the methionine can be used again.
Folate Balance
Vitamin B12 is the only acceptor of methyl-THF. There is also only one acceptor for methylB12 which is homocysteine in a reaction catalyzed by homocysteine methyltransferase.
This is important because a defect in homocysteine methyltransferase or a deficiency of
B12 can lead to a methyl-trap of THF and a subsequent deficiency. Thus, a deficiency in
B12 can generate a large pool of methyl-THF that is unable to undergo reactions and will
mimic folate deficiency. Another form of THF, formyl-THF or folinic acid) results from
oxidation of methylene-THF or is formed from formate donating formyl group to THF.
Methotrexate
An anti-folate
Structure similarity of folic acid (left) and
methotrexate (right)
Methotrexate competitively and irreversibly inhibits dihydrofolate reductase (DHFR), an enzyme
in tetrahydrofolate synthesis. The prototypic antifol DHFR inhibitor is a 4-amino–substituted
pterin compound. The substitution of an amino moiety for 4-hydroxyl results in a folate
analogue with several thousand-fold increase in affinity for DHFR. The Ki of MTX for DHFR is
below 10-10 M, well below the Km of the natural substrate, dihydrofolate, which is in the
micromolar range. It is this remarkable increase in affinity compared with the natural substrate
that resulted in MTX being considered a “stoichiometric inhibitor” of DHFR. Folic acid is
needed for the de novo synthesis of thymidine (DNA), for purine base synthesis, so all purine
synthesis will be inhibited. Methotrexate, therefore, inhibits the synthesis of DNA, RNA,
thymidylates, and proteins.
Methotrexate acts specifically during DNA and RNA synthesis, and thus it is cytotoxic during the S-phase of
the cell cycle. It therefore has a greater toxic effect on rapidly dividing cells which replicate their DNA more
frequently, and thus inhibits the growth and proliferation of these non-cancerous cells as well as causing the
side effects listed below.
B12 - Cobalamin
Reductive elimination
Oxidative addition
Monsanto Acetic Acid Catalysis
(organometallic catalysis in industry)
The Major Pterin Players
Neopterin: biochemical precursor; immune response marker
Folate/Folic acid: One carbon (C1) pool
Biopterin: redox cofactor for (1) aromatic aminoacid hydroxlases (neurotransmitters)
(2) NOS (nitric oxide synthase), regulator, signaling
Methanoopterin: C1 chemistry (methyl transfer) methanogenesis
Molybdopterin: O atom chemistry
Synthetic pterins: methotrexate
anti-folates
Folate  C1 chemistry
How they’re related (or not)
Biopterin  PAH cofactor
NOS cofactor
Neopterin
GTP
Guanosine triphosphate
Methopterin
X
Molybdopterin  Mo / W enzymes
…molybdopterin biosynthesis proceeds via
a pathway unlike that of any known
pteridine biosynthetic pathway.
Folates
Folic acid (vitamin B9), tetrahydrofolate (cofactor), dihydrofolate,
Etymology: Folate and folic acid derive from the Latin word folium, "leaf”
C1 chemistry: methylation
(a)Homocysteine to methionine; (b) uracil to thymine; (c) purine synthesis
• Nucleic acid metabolism involve the addition of C1 units.
• C1 units are derived from the breakdown of serine and
glycine and carried by the coenzyme tetrahydrofolate (H4F),
which is synthesized from folic acid in the diet.
• These C1 fragments can be oxidized and reduced while
bound to the H4F coenzyme, so they may be inserted into the
receiving molecules as formyl groups (most oxidized),
hydroxymethyl or methyl groups (most reduced).
[search Expasy: tetrahydrofolate]
• Methylene-THF (CH2FH4) is formed from THF by the
addition of methylene groups from one of three carbon
donors: formaldehyde, serine, or glycine. Methyl
tetrahydrofolate (CH3-THF) can be made from methylene-THF
by reduction of the methylene group with NADPH. Histidine
can donate a single carbon to THF to form methenyl-THF.
Folates
Folic acid (vitamin B9), tetrahydrofolate (cofactor), dihydrofolate,
Etimology: Folate and folic acid derive from the Latin word folium, "leaf”
C1 chemistry: methylation
(a)Homocysteine to methionine; (b) uracil to thymine; (c) purine synthesis
Health
Lack of dietary folic acid leads to folate deficiency (FD). Resulting health problems:
(1)neural tube defects in developing embryos. Neural tube closure takes place during early embryogenesis
and requires interactions between genetic and environmental factors. Failure of neural tube closure is a
common congenital malformation that results in morbidity and mortality. A major clinical achievement has
been the use of folic acid supplements, which prevents ~50–75% of cases of neural tube defects. However,
the mechanism underlying the beneficial effects of folic acid is far from clear. Biochemical, genetic and
epidemiological observations have led to the development of the methylation hypothesis, which suggests
that folic acid prevents neural tube defects by stimulating cellular methylation reactions.
(2) homocysteine accumulation as a result of one carbon metabolism mechanism being impaired.
(3) DNA synthesis and repair are impaired and this could lead to cancer development.
A 2003 opinion article in the New York Times: Adding folic acid and micronutrients to the food supply of
developing countries would have a greater impact than any other single action in improving world health.
Methanopterin
A cousin to the folates, does C1 chemistry.
A cofactor in methanogenesis, a bacterial process of methane production in respiration.
Differs from folates in that phenyl group lacks a carbonyl, so is harder to reduce.
[search Espasy: methanopterin]
Biopterin
THB site
Redox cofactor as tetrahdrobiopterin
First discovered as cofactor (S. Kaufman) for PAH, phenylalanine hydroxylase;
role = activation of O2 for para-hydroxylation of phenyl group
Later revealed as cofactor for analogous hydroxylations of tyrosine (to L-DOPA), tryptophan
(to 5-hydroxy-tryptophan ).
The class of enzymes is called the aromatic amino acid hydroxylases.
These are precursors to neurotransmitters serotonin, melatonin, dopamine,
norepinephrine (noradrenaline), epinephrine (adrenaline).
Most recently identified as cofactor for different substrate formation, NO nitric oxide, also
having neurotransmitter function. Enzyme is NOS, nitric oxide synthase.
Biopterin
THB site
Health and disease issues
PKU, phenylketonuria, excess phenylalanine—PKU can result in severe brain damage,
including symptoms of mental retardation, microcenphaly, speech impediments such as
stuttering, slurring, and lisps, seizures or convulsions, and behavioral abnormalities.
When this condition was first recognized in the 1930's, a significant proportion of all long
term patients in mental institutions proved to be undiagnosed phenylketonuriacs.
Approximately one person in 45 American whites is a carrier for a defective
phenylalanine hydroxylase gene. These mutations are particularly common in people of
Celtic origin, but are less frequent in Eastern Europe.
Less frequent is THBD, tetrahydrobiopterin deficiency.
Rare genetic condition: 450 cases, causing only 1-2% of PKU patients
Synthetic tetrahydrobiopterin, developed by BioMarin under the brand name Kuvan and
approved by FDA on December 13, 2007, is a synthetic preparation of the
dihydrochloride salt of the substance, used in the treatment of PKU
Neopterin
Biosynthetic precursor in pterin pool;
for biopterin cofactor of the aromatic amino acid hydroxylases and nitric oxide synthase.
[search Espasy: neopterin]
Health and disease issues
• Induced synthesis from GTP by macrophages upon stimulation with the cytokine interferon-gamma and
is indicative of a pro-inflammatory immune status.
• Neopterin serves as a marker of cellular immune system activation (measured in body fluids: blood
serum, cerebrospinal fluid or urine) .
• Why is it formed? A result of activation of GTP cyclohydrolase I; so an excretory product of GTP
metabolism. Catabolism to BH4 not induced so neopterin builds up.
• High neopterin production is associated with increased production of reactive oxygen species, neopterin
concentrations also allow to estimate the extent of oxidative stress elicited by the immune system.
Increased neopterin production is found in, but not limited to, the following diseases:
* viral infections including human immunodeficiency virus (HIV), hepatitis B and Hepatitis C
* bacterial infections by intracellular bacteria: borrelia (Lyme Disease) and mycobacterium tuberculosis
* parasites such as malaria
* autoimmune diseases such as rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE)
* malignant tumor diseases
* allograft rejection episodes.
Anti-Folates
A number of drugs interfere with the
biosynthesis of folic acid and THF.
Among them are the dihydrofolate
reductase inhibitors such as trimethoprim,
pyrimethamine and methotrexate; the
sulfonamides (competitive inhibitors of
para-aminobenzoic acid in the reactions of
dihydropteroate synthetase).
Lower doses of methotrexate have been shown to be very effective for the management of
rheumatoid arthritis, Crohn's disease, and psoriasis. For the treatment of rheumatoid
arthritis, patients should supplement their diet with folate. In these cases inhibition of
dihydrofolate reductase (DHFR) is not thought to be the main mechanism, but rather the
inhibition of enzymes involved in purine metabolism, leading to accumulation of adenosine,
or the inhibition of T cell activation and suppression of intercellular adhesion molecule
expression by T cells.