Tetrahydrofolate Lecture Sep 6 Flashcards
Rank the following from most reduced carbon to moves oxidized carbon:
Carboxylic acid, methyl, alcohol, aldehyde
Methyl (most reduced)
Alcohol
Aldehyde
Carboxylic acid (most oxidized)
What are five sources of one-carbon units?
What molecules comprise the one-carbon pool?
Five sources:
- Serine
- Glycine
- Histidine
- Formaldehyde
- Formate
The one-carbon pool: (bound to FH4)
- Formyl
- Methylene
- Methyl
What is the vitamin precursor for FH4?
Where is it found in the diet?
Folate
Found in green leafy vegetables, liver, legumes, yeast, and fortified flour
What happens to folate after it has been eaten before it becomes FH4?
It has a poly-glutamate “tail” that is digested in the gut down to monoglutamate.
Folate is then reduced tot he N5-Methyl FH4 in the intestinal epithelial cells
Then it’s poly glutamated again in the liver
Folate deficiency before and during pregnancy is associated with what condition?
Neural tube defects like spina bifida.
Patients who ar eplanning to get pregnant should be advised to take daily folate for a month before and three months after getting pregnant
What mutation would cause a functional polar deficiency despite adequate folate in the diet?
Hereditary folate malabsorption:
This is an inherited mutation in the proton coupled folate transporter (PCFT; gene SCL46A1)
This means the folate doesn’t get absorbed through the gut.
What enzyme reduces dietary folate to FH4? THrough what steps?
- Folate is reduced to dihydrofolate (FH2) and again to tetrahydrofolate (FH4) by the enzyme dihydrofolate reductase (DHFR)
DHFR will reduce dietary folate to FH4 and it will also recycle oxidized folate (FH2) back down to FH4
DHFR is an important drug target. What are three drugs that target DHFR?
Methotrexate–for cancer and RA
Trimethoprim–antibacterial
Pyrimethamine–antimalarial
What are the 4 forms of tetrahydrofolate?
10-Formyl FH4
5,10-Methylene FH4
5,10-Methenyl FH4
5-Methyl FH4
What single carbon molecule does tryptophan donate to FH4?
Formate
Where does formate bind to FH4?
What is the result?
It adds to the 10-N
This results in N10-formyl FH4
What is the order of FH4 forms from most oxidized to most reduced.
- Formyl adds on at the 10N location, yielding the most oxidized form: N10-formyl FH4
- N10-formyl FH4 is reduced to 5,10-Methenyl FH4
- Next is 5.10-Methylene FH4
- The most reduces form is 5-Methyl FH4
What is the methyl trap?
While most reduction steps that form the different FH4 varieties are reversible, the step creating 5-methyl FH4 is not. So once FH4 becomes completely reduces, it cannot become more oxidized. This “traps” it as methyl FH4.
How does histidine donate single carbon molecules to FH4?
Histidine degrades and donates an HC=NH group to FH4, yielding 5-Formimino FH4 (also called FIGLU).
FIGLU then is deaminated to form 5.10-Methenyl FH4.
Then the rest of the FH4 forms can be generated.
How does serine donate a single carbon molecule to FH4?
What is the product of this reaction?
When serine is acted on by serine hydroxymethyl transferase (in the presence of PLP) it trasfers serine’s side chain to FH4 to make glycine.
This forms 5,10-methlenetetrahydrofolate, which then reorganizes to 5,10-Methylene FH4
Which amino acid is the most important contributor to the one carbon pool?
At what level of FH4 forms does it enter?
Serine
It enters at 5,10-methylene FH4
After the carbon molecules are donated to FH4, what are some reaction products it can produce (i.e. donate carbon groups to)?
FH4 can donate to make thymidine nucleotide (for DNA), purine bases, methionine, and S-andenosyl methionine
When oxidized carbon is donated from 5,10-methylene FH4 to dUMP, what are the two products?
thymidine monophosphate (TMP) and dihydrofolate (FH2), which is then reduced to FH4 with dihydrofolate reductase
What enzyme converts dUMP to dTMP?
Thymidylate synthase (TS)
It uses methylene FH4 and oxidizes it to FH2
What does dietary folate deficiency result in?
Megaloblastic anema
the RBCs cannot synthesize enough DNA to replicate their chromosomes so they don’t divide and grow very large.
So you get a low blood cell count and the RBCs you do have are enlarged.
WHy do methotrexate and 5-fluorouracil work as cancer drugs?
Cancer cells proliferate rapidly, so they need rapid DNA synthesis.
Without a well functioning folate pathway, this DNA synthesis cannot occur because there isn’t enough dTMP
5-Fluorouracil is a uracil analog that blocks thymidylate synthase, so there is no conversion of dUMP to dTMP.
Methotrexate is a dihydrofolate analog that will competitively inhibit DHFR.
What are the steps for the donation of REDUCED carbon?
- N5-Methyl FH4 donates the methyl group to cobalamin to make methylcobalamin.
- Methylcobalamin donates the methyl group to homocysteine to make methionine
- Methionine joins an adenosine base to form S-adenosyl methionine (SAM)
- SAM is then the actual methyl donator for diosynthetic reactions, regenerating homocysteine.
What reactions are SAMs commonly used in?
Donation of methyl groups to NT precursors to make the active neurotransmitters lke epinephrine and melatonin.
Also used for some nucleotides and phospholipids.
5N methyl FH4 can only donate its methyl group to one thing. What is that?
It can only donate to cobalamin to form methylcobalamin.
Methylcobalamin participates in only one reaction. What is it?
Donation of a methyl to homocysteine to make methionine.
Adding an adenosine base onto cobalamin yields adenosylcobalamin. What reaction does this catalyze?
It is important in converting branched chain AAs, branched chain fatty acids, and odd length fatty acids into useful fuel.
It converts methylmalonyl CoA (a branchd fatty acyl CoA) into succinyl CoA (a linear fatty acyl CoA), which can enter the TCA cycle.
The cobalt in the cobalamine complex can bind 1 of two things. What are they?
Either a methyl group (methylcobalamin which then donates to homocystein to make cysteine)
or an adenine nucleotide (making adenosylcobalamin which converts methylmalonyl CoA to succinyl CoA)
What happens to cobalamin (vitamin B12) after it’s ingested?
- Cobalamin binds to R-binder proteins secreted in the stomach.
- After the cobalamin-R-binder protein passes through into the intestine, the R-protein starts to be digested by pancreatic proteases.
- This allows the cobalamin to bind another protein, instrinsic factor (which was also secreted in the stomach)
- The intrinsic factor-cobalamin complex is taken up by intestinal epithelial cells
- Transcobalamin II binds the intrinsic factor/cobalamin and transports it through the blood either for use in tissues or for storage in the liver.
Cobalamin circulating in the blood is bound to what protein?
Transcobalamin II protein
What organ stores cobalamin? In a complex with what protein?
Most of it is stored in the liber in complex with cubillin
WHat does cobalamin deficiency result in?
pernicious anemia
which is basically megaloblastic anemia plus neuropathy
What are 4 potential causes of cobalamin deficiency?
- dietary deficiency
- loss of function in the instrinsic factor (many cases are caused by autoimmune destruction of parietal cells)
- loss of transcoalamin II function
- loss of cubillin function
Cobalamin is used for two reactions in the body. WHat are they?
- Adenosyl cobalamin is a cofactor for methylmalonyl CoA mutate, which converts methylmalonyl to succinyl CoA
- Methylcobalamin is a cofactor for methionine synthase, which catalyzes the transfer of methyl from methylcobalamin to homocystein to make methionine.
How is cobalamin (either as adenosylcobalamin or methylcobalamin) regenerated after their respective reactions?
Adenosylcobalamin is not consumed in the reaction.
Methylcobalamin is regenerated by accepting a methyl from fully reduced N5-methyl FH4
Mutations in methionine synthase will result in what?
hyperhomocyteinemia
this is the buildup of homocysteine because it can’t be converted to methionine
it’s linked to cardiovascular and neurological problems
What are the three ways hyperhomocysteinemia can arrise?
- Cobalamin deficiency
- Methionine synthase deficiency
- Vitamin B6 (pyridoxal phosphate) deficiency-because PLP is needed for amine transfer reactions
Where do the two cobalamin-requiring reactions occur within the cell?
The methionine synthase reaction occurs in the cytosol
The methylmalonyl CoA reaction occurs in the mitochondrial matrix
Why do you get folate deficiency symptoms with a cobalamin deficiency?
The ONLY reaction that 5-methl FH4 can take place in is losing its methyl to cobalamin to form methylcobalamin.
In a dietary or functional deficiency of cobalamin, folate becomes trapped as N5-methyl FH4 because it has no cobalamin to give its methyl to.
Thus, the folate can’t participate in other one carbon transfers so you develop folate deficiency symptoms even though there is plenty of FH4 in the blood–it’s just in a useless form
Why cobalamin deficiencies have neurological symptoms when folate deficiencies don’t?
Cobalamin has a role in transferring methyl groups to SAMs, which then transfer the methyl groups to neurotransmitters.
