Methylation Reactions

What are some characteristics of folate? Why is it important?

• vitamin B9

• It is a necessary dietary vitamin needed for DNA synthesis and homocysteine conversion; it is the precursor of tetrahydrofolate.

• Folate is found in leafy veggies and fortified foods like bread and whatnot.

• Folate can be converted to tetrahydrofolate (THF). The location of the 1-C determines the name and function of the THF:

  • R-COOH is formyl,

  • R-CH= is methenyl, and

  • R-CH2 is methylene.

• THF is a carbon donator, acting as a cofactor for nucleic acid and amino acid synthesis.

Describe the reaction of folate with DHFR. What is the product? What is required?

Folate is converted into DHF by DHFR (dihydrofolate reductase), which requires NADPH.

Describe the reaction of DHF with DHFR. What is the product? What is required?

DHFR reacts again with DHF (dihydrofolate) to make THF, also requiring NADPH

What is methotrexate? What is its function and how does it accomplish that?

DHFR is inhibited by methotrexate. It has a similar structure to folic acid (NOTE THE DIFFERENCES!!!)

It inhibits DNA synthesis from TMP by preventing the conversion of DHF to THF. THF would be converted to deoxyuridine monophosphate (dUMP), which is converted to thymidine monophosphate (TMP) used for DNA.

In summary, it is a cancer drug that inhibits DNA synthesis.

What is 5-FU (fluorouracil)? What is its function and how does it accomplish that?

It is one of the first chemotherapy drugs with a similar structure to uracil. This inhibits thymidylate synthase, preventing formation of TMP for use in DNA synthesis.

Describe the reaction of THF with SHMT. What is the product? What is required?

Serine Hydroxymethyltransferase (SHMT) converts THF into N5, N10 methylene THF.

This reaction uses serine and converts it into glycine (functions as a hydroxy donor).

This reaction requires pyridoxal phosphate (B6).

Describe the reaction of N5N10 methylene THF with thymidylate synthase? What on earth does this achieve? What could possibly be required?

N10 methylene THF can be converted back into DHF by thymidylate synthase. This requires dUMP, which converts it into dTMP (remember the DNA synthesis path mentioned previously in your mind). Also requires FADH2, which is oxidized into FAD+. Thymidylate synthase is inhibited by 5-FU (fluorouracil).

Describe the reaction of N5N10 methylene THF with MTHFR. In what conditions does this reaction take place? What is the product? What is required?

If DNA synthesis (as from dTMP) is not needed, N5, N10 methylene THF is turned into 5-methyl THF by methylene THF reductase (MTHFR). This reaction requires NADPH and FAD+, producing FADH2 and NADP+. This reaction is INHIBITED by DHF (from early in the pathway) and SAM (S-adenosylmethionine)

Describe the reaction of 5-methyl THF with methionine synthase. What is another name for this enzyme? What is the product? What is required?

5-methyl THF is converted back into THF by methionine synthase (AKA homocysteine methyltransferase). This requires vitamin B12. This reaction uses homocysteine and converts it into methionine (obviously).

Describe the reaction of methionine with methionine adenoyltransferase. What is the product? What is required?

Methionine is converted into S-adenosylmethionine (SAM) by methionine adenosyltransferase (this can inhibit MTHFR as stated by your mind). SAM can be converted to homocysteine by a few sinister unimportant processes. This whole ordeal with homocysteine and methionine is called the activated methyl cycle.

Describe the reaction of N5N10 methylene THF with MTHFD. When does this occur? What is the product? What is required?

if there are no other ambitions for N5N10 methylene THF, it gets converted into N5N10 methenYL THF by MTHFD (methylene THF dehydrogenase). This requires NADPH and makes NADP+

Describe the reaction of N5N10 methenYL THF with MTHFC. What is another name for this enzyme?When does this occur? What is the product? What is required?

N5N10 metheYL THF is converted into N10-formyl THF by methenyl THF cyclase/cyclohydratase. This requires H2O. The formyl can be used for purine synthesis.

What are the three main molecules of the folate cycle?

What is homocystineuria? What are some symptoms?

Homocystinuria occurs when levels of homocysteine are high. Some symptoms include deep vein thrombosis, stroke, atherosclerosis, marfan-like habitus, mental retardation, and joint contractures.

What are the biochemical consequences of folate deficiency? How is it overcome?

Since folate is unavailable for some reason, the folate cycle is unable to continue. The active folate pool diminishes and leads to an inability to perform DNA and RNA cell division. This leads to megaloblastic anemia, in which RBCs are unable to grow or divide, leading to lack of replication and unusually large blood cells. This can be overcome by folate adminstration.

What are the biochemical consequences of B12 deficiency? How is it overcome?

B12 is responsible for the mobilization of the storage pool (5-methyl THF) into THF to be made into active folate for real shenanigans like DNA synthesis. B12 is required for methionine synthase to make THF, and therefore lack of B12 leaves all that sweet methyl THF stuck in the storage pool (90% of THF is stored there at all times). Because there is no other direction of escape from this storage other than through MS, this leads to a lack of THF and therefore a lack of DNA synthesis, and megaloblastic anemia similarly to folate deficiency.

How exactly does folate deficiency differ from B12 deficiency? What treatment methods are commonly used?

Other differentiating factors of B12 deficiency is progressive peripheral neuropathy (numbness, weakness, shooting pains), and issues with METHYL MALONYL COA MUTASE (also requires B12). This will leave METHYL MALONIC ACID in the URINE.

What treatment methods are commonly used for B12/folate deficiency

Since both result in megaloblastic anemia, the treatment option for this sign is to provide BOTH B12 and folate. Providing folate can be sufficient treatment for both problems, but the UNDERLYING PROBLEM OF B12 DEFICIENCY WILL REMAIN and you are still suffering until you get that B12. 1

What is transmethylation?

The transfer of a -CH3 group from ACTIVE methionine to an acceptor. Methionine has to be ACTIVE (to S-adenosylmethionine) to donate the methyl like I just said. The sulfur makes it active (like in CoA).

Describe the SAM cycle.

Requires 2 ATP essentially. Methyltransferases are involved in the transfer of methyl groups.

What is a catechole?

A molecule with a 3,4 dihydroxy phenyl ring. An example of this is DOPA, dopamine, norepinephrine, and epinephrine

What is the rate limiting enzyme of epinephrine synthesis?

Hydroylase.

This transforms tyrosine into DOPA.

What is the role of epinephrine?

A vasoconstrictor that can be added to local anesthetics to keep the agent in circulation.