5/6 bchm2

Metabolism of Sulfur-Containing Amino Acids

Review of Amino Acid Metabolism

The lecture starts by discussing the metabolism of sulfur-containing amino acids, specifically how the sulfur group is metabolized.
A review of previous material from before midterm three covers the metabolism of alanine, serine, and glycine, focusing on their conversion into pyruvate.
These conversions are really focusing on the reactions that can produce either pyruvate directly, or can be converted to pyruvate by other means.

Alanine Metabolism

Alanine's conversion to pyruvate is the simplest because pyruvate is the alpha-keto acid form of alanine.
The amino transfer from alanine to alpha-ketoglutarate generates pyruvate.

Serine Metabolism

Serine's conversion involves a dehydration reaction driven by the serine dehydratase enzyme, which is PLP-dependent.
This dehydration reaction produces an enamine, which rearranges to an imine and then undergoes spontaneous hydrolysis in the presence of water, releasing ammonia.

Glycine Metabolism

Glycine's metabolism is more complex, as pyruvate is a three-carbon molecule, and glycine needs an intermediate for conversion.
Stoichiometrically, two glycines are converted into one serine and one $CO_2$ . This accounts for all four carbons initially present in the two glycine molecules.
The serine produced is then converted to pyruvate through the serine dehydratase reaction.
Overall stoichiometry: 2 glycines → 1 pyruvate + $CO_2$ .

Tetrahydrofolate

The cofactor tetrahydrofolate is crucial in this process.
Two separate enzymatic reactions occur involving glycines:
- One glycine is directly converted to serine via the serine hydroxymethyltransferase enzyme, which transfers a carbon unit from methylene tetrahydrofolate to make serine.
- The second glycine is used in the glycine cleavage enzyme, which generates methylene tetrahydrofolate.
The glycine cleavage enzyme converts tetrahydrofolate to methylene tetrahydrofolate, an oxidative decarboxylation reaction that also requires the redox cofactor $NAD^+$ , which is converted to $NADH$ .
The glycine cleavage enzyme resembles the pyruvate dehydrogenase complex and alpha-ketoglutarate dehydrogenase complex, which are complex enzymes made of multiple parts catalyzing oxidative decarboxylation reactions.
Like pyruvate dehydrogenase, glycine cleavage uses lipoylisine cofactors and three redox cofactors.
Glycine cleavage differs by using PLP instead of TPP as the initial cofactor due to the amino acid substrate, while the other complexes use alpha-keto acids.
Instead of transferring a two-carbon unit, glycine cleavage transfers a one-carbon unit (methylene group) to tetrahydrofolate.

Serine Hydroxymethyltransferase

The reaction for serine hydroxymethyltransferase begins with the formation of an external aldimine with glycine.
The initial step involves deprotonation, creating a carbanion intermediate.
This carbanion then reacts with methylene tetrahydrofolate, adding a $CH_2OH$ group to convert glycine to serine.
A water molecule then comes in, and the hydroxyl of the water molecule replaces the carbon nitrogen bond with a carbon-oxygen bond through an SN2 like reaction (more accurately, displacement of the amino acid with a lysine side chain).
The enzyme's lysine side chain forms an internal aldimine, releasing serine; the reaction is reversible.
This reversibility allows interconversion of glycine and serine metabolism. Serine can then enter the serine dehydratase reaction to be converted to pyruvate.
These two enzyme reactions account for the conversion of two-carbon glycine into the three-carbon serine intermediate.

General Amino Acid Oxidation

Amino acid oxidation typically involves amino group transfer first (except for serine and threonine, which can undergo direct dehydration), followed by oxidation of the carbon skeleton through the citric acid cycle.
Carbon skeletons enter the citric acid cycle at different entry points.
The enzyme that releases serine is the lysine side chain in the active site of the protein, which normally forms an internal aldimine.

Ketogenic vs. Glucogenic Amino Acids

Amino acids that are converted to acetyl CoA directly are ketogenic because they cannot be used for glucose metabolism.
Amino acids that are converted to pyruvate or other citric acid cycle intermediates are glucogenic because these intermediates can be used for gluconeogenesis.
Some amino acids are both ketogenic and glucogenic if parts of their skeletons are converted to acetate and other parts to intermediates like pyruvate.

Aminotransferases and Glutamate Dehydrogenase

PLP-dependent aminotransferase enzymes transfer the amino group from an amino acid to alpha-ketoglutarate, producing an alpha-keto acid and glutamate.
Glutaminase cleaves glutamate through an oxidative NAD+-dependent reaction, releasing free ammonia and regenerating alpha-ketoglutarate.
Amino acid carbon skeletons are either glucogenic or ketogenic.
Glycine, being a small two-carbon amino acid, requires two molecules to be converted into pyruvate, using PLP and tetrahydrofolate as cofactors and producing $CO_2$ and two ammonia molecules.
The reaction pathway for glycine also generates $NADH$ , which can be used in oxidative phosphorylation.

Tetrahydrofolate in One-Carbon Metabolism

Tetrahydrofolate is a central cofactor in one-carbon metabolism.
The lecture will continue discussing this in the context of sulfur-containing amino acids.

Methionine Metabolism

Methionine metabolism is used not only for protein biosynthesis, but also for other metabolic processes.
Methionine contains a sulfur atom and a single isolated methyl group, bridging both single-carbon and sulfur amino acid metabolism.
The reactions discussed include both breakdown and recycling of methionine.
A key intermediate in the methionine breakdown pathway is S-adenosylmethionine (SAM), which is produced from methionine using ATP.
The reaction forming SAM is unique because all three phosphates are cleaved from ATP in the same reaction with the the triphosphate isn't directly released from the enzyme, but it actually gets cleaved after the three phosphates are kicked off of the ATP.
Hydrolysis of the triphosphate into pyrophosphate and inorganic phosphate makes the reaction irreversible.
The sulfonium ion in SAM allows it to easily transfer its methyl, adenylate, or amino acid side chain groups like we see in Polyamine biosynthesis.
The most common transfer is of the methyl group.

Function of SAM

SAM (also known as adenosylmethionine or ADOMET) is a major source of methyl groups in central metabolism.
It is estimated to be one of the most abundant cofactors in cells after ATP.
When SAM transfers a methyl group, it becomes S-adenosylhomocysteine (SAH).

Examples of SAM-dependent Methylation

Carnitine biosynthesis: Three methyl groups on carnitine are introduced by SAM.
Creatine biosynthesis: The methyl group on creatine is introduced by SAM.
Epinephrine synthesis: SAM methylates norepinephrine to form epinephrine, a key hormone in the fight-or-flight response.
DNA methylation: Methyl groups on cytosine residues in DNA are added by SAM and serve as epigenetic marks.
Protein methylation: Histone proteins are methylated by SAM-dependent enzymes, influencing transcriptional regulation.

Recycling of Methionine

After methyl group transfer and the creation of SAH, the next step is to hydrolyze SAH, replacing the carbon-sulfur bond with a carbon-oxygen bond, yielding adenosine and homocysteine.
Homocysteine can be methylated again using N5-methyl tetrahydrofolate and coenzyme B12 to regenerate methionine in a radical-based reaction.
The B12-dependent enzymes spontaneously generate a radical through the cleavage of vitamin B12.
This process constitutes an amino acid recycling mechanism. SAM radical reaction will generate that five prime adenosyl radical.
In general, all radical-based chemistry involving B12-dependent enzymes have in common the generation of adenosyl radicals.
Radical-based mechanisms facilitate energetically unfavorable reactions like methyl transfer to homocysteine.
A cyclic set of reactions interconverts methionine to SAM and then to homocysteine using methyl tetrahydrofolate.

Methionine Oxidation

The process of oxidizing methionine starts with same steps that we just saw.
The key intermediate in both oxidation and recycling of methionine is homocysteine.
Methionine is a glucogenic amino acid, converted to succinyl CoA.
To make succinyl CoA, methionine is converted to propionyl CoA first.

Breakdown Pathway of Methionine to Succinyl CoA

The initial steps in methionine breakdown mirror those in the recycling process.
Homocysteine reacts with serine, in a PLP dependent reaction, to generate cystathionine due to the serine reacting with PLP, similar to serine dehydratase.
Then, cystathionine lyase cleaves cystathionine (another PLP dependent step) to form α-ketobutyrate, cysteine, and ammonia.
This is similar to the dehydration reactions we have seen before. We are losing $NH_4$ the same was has how we do it in those dehydration reactions for breakdown of for breakdown of serine.
$NH_4$ isn't generated through dehydration reaction, but it's the similar mechanism as what we saw.
Humans require methionine and can biosynthesize with cysteine through this cystathione reaction.
Cystathionine is an important link between methionine degradation and cysteine biosynthesis.
The overall reaction mechanism of said lyase is a complex involving transfer reactions. During the reaction, an enamine is cleaved which them undergoes spontaneous keto enotomerization.
The enamine undergoes rearrangement to spontaneous hydrolysis similar to serine.
The next step in the pathway is to convert α-ketobutyrate to propionyl CoA using an α-keto acid dehydrogenase complex, which is analogous to pyruvate dehydrogenase complex.
After this step we go through a series of reactions to convert propionyl CoA to succinyl CoA through a series of known reactions
The cleavage reaction in the α-keto acid dehydrogenase complex is an oxidative decarboxylation producing $CO_2$ and using TPP, lipoamide, FAD+ cofactors along with coenzyme A to generate propionyl-CoA. Remember that because it's propionyl-CoA gets converted to succinyl-CoA, you need to use that b12 rearrangement again

Cysteine and Sulfur Metabolism

Methionine and cysteine both require the recycling of sulfur atoms.
Breakdown of cysteine involves an enzyme that combines cysteine and homoserine to become cystanthionine, so we are just using homoserine in the place of serine, and breaking it down on the opposite end.
Sulfur metabolism involves exchange of sulfur atoms between amino acids without the release of free sulfur. Assimilation of sulfur is typically done by microorganisms.
Methyl group is transferred to methionine, which then gets transferred to all kinds of the things through acid endosymethionine.
SAM then gets converted to homocysteine through that pathway, and then the homocysteine either gets recycled using that 5 methylphyl tetrahydrofolate.
Eventually, that acid methionine is converted to homocysteine through that pathway, and then the homocysteine either gets recycled using that 5 methylphyl tetrahydrofolate through methyl transfer, or it's broken down to make succinyl CoA, and that succinyl CoA enters our citric acid cycle.
For full oxidation it generated the same products for full oxidation if as if we were breaking down propionyl CoA.
In order to get converted to succinyl CoA, first gets converted well, first, the homocysteine. Eventually, that homocysteine gets converted to alpha ketobutyrate.
Alpha ketobutyrate undergoes oxidative dehydrogenase which is the same type of complex as pyruvate dehydrogenase complex.
We can think of enzymes being organized into three protein components. The first is an enzyme that contains the TPP cofactor and initially reacts with the alpha keto acid. There is an enzyme that contains the swing arm, which is responsible for transferring the product onto the lipo lysine. Finally, there's an enzyme that contains FAD, is involved in reducing lipo-lysine, and getting re re-oxidized to use $NAD^+$ .