|
Amino acid metabolism Amino acid metabolism Degradation Because of the multitude of pathways, only a selected few are presented here demonstrating basic principles found in all other metabolic pathways of nitrogenous compounds. Amino acids serve as precursors for lipids, carbohydrates, and nucleic acids including ribonucleotides used as cosubstrates and coenzymes in the production of energy (ATP, NAD, FAD, CoA). Following the metabolic fate of carbon atoms of dietary amino acids, they can be traced to all major metabolic intermediates because of the close interaction of amino acid metabolism with both the citric acid cycle and glycolysis/gluconeogenesis. These intermediates containing carbons from dietary amino acids include pyruvate, acetyl-CoA and acetoacetyl-CoA (ketone bodies), and the citric acid cycle intermediates a-ketoglutarate, succinyl-CoA (heme synthesis), fumarate, malate, and oxaloacetate. Amino acid metabolism is 'separated' into pathways according to the different length of carbon structures involved. These are referred to as the C3, C4, and C5 families of amino acids. which produce common end products during catabolism. The C3 family includes alanine, serine (glycine), and cysteine, all of which are degraded to pyruvate. They are glucogenic amino acids because they can directly be utilized by the liver gluconeogensis (except their amino groups which are excreted as urea). The C4 family of amino acids includes aspartate and asparagine, which are degraded to oxaloacetate and are closely linked to glutamate and alpha-ketoglutarate interconversion by amino transferases. The C4 amino acid threonine has a separate pathway leading to pyruvate and is a glucogenic amino acid. The C5 family of amino acids includes glutamine, proline, arginine, and histidine, all of which are converted ultimately to glutamate, which is deaminated to alpha-ketoglutarate. The non-polar C4 amino acids methionine, isoleucine and valine are precursors for the synthesis of odd numbered fatty acids via the intermediate propionyl-CoA (a C3 acyl-CoA). Propionyl-CoA can be reused for succinyl-CoA (C4) synthesis (carboxylation) which in turn serves as heme precursor. The non polar amino acid leucine, however, undergoes a more complex degradation pathway, including a decarboxylation-carboxylation detour leading to the formation of acetyl-CoA and acetoacetyl-CoA (a ketone body). Metabolism of aromatic
amino acids Aromatic amino acids include phenylalanine, tyrosine and tryptophan. Phe and Tyr are closely related. They contain a benzene ring which is hydroxylated in tyrosine. Tyrosine is synthesized directly from the essential amino acid phenylalanine. Tryptophan contains a conjugated indole ring and its metabolism is linked to that of vitamin B (niacin C00253). These metabolic relations give rise to an intricate nutritional dependence. For example, a high level of dietary tyrosine relieves the need for essential phenylalanine. Also, metabolic disorders like the impairment of synthesizing tyrosine from phenylalanine makes the former an essential amino acid. This lack of amino acid biosynthetic pathways in humans is the cause of many diseases associated to malnutrition. Pellagra is a vitamin deficiency syndrome caused by an inadequate supply of niacin (vitamin B) because of problems in the pathway leading from tryptophan to niacin synthesis. Vitamin C (C00072), which is a necessary coenzyme in tyrosine metabolism, or vitamin B6 (C00250), which is required for tryptophan metabolism, cause deficiencies in the metabolism of aromatic amino acids. Degradation of aromatic ring structures is mostly performed in liver, also many specialized cells can use the benzene and indole rings for the synthesis of more complex, biologically important molecules such as heme, pigments, and hormones. Phenylalanine Phenylalanine is first converted to tyrosine by the addition of a hydroxyl unit. This reaction is catalyzed by phenylalanine hydroxylase (EC 1.14.16.1). This is a liver specific enzyme and belongs to the group of monooxygenases. The reaction requires molecular oxygen, NADPH, and the coenzyme tetra-hydrobiopterine (C00272). Note that the hydroxylation of the benzene ring is used to destabilize it preparing the ring structure for breakup. The cofactor hydrobiopterine is oxidized during tyrosine formation. It is converted back to its reduced form by dihydrobiopterin reductase (EC 1.6.99.7). The reaction of phenylalanine hydroxylase itself is irreversible. Tryosine Tyrosine degradation is a liver resident process. It starts out with the transfer of its amino group to alpha-ketoglutarate by tyrosine-glutamate aminotransferase (EC 2.6.1.5). This enzyme is specific for both tyrosine and phenylalanine. Other L-tyrosine accepting transaminases are less substrate specific, including aspartate transaminase (EC 2.6.1.1) which acts on aspartate, tyrosine, phenylalanine, and tryptophan. This latter enzyme isoform is obtained from aromatic-amino-acid transaminase (EC 2.6.1.57) by controlled proteolysis. The degradation intermediate of this transaminase reaction is 4-hydroxy-phenylpyruvate (C01179) which in turn is oxidized in the presence of vitamin C to homogentisic acid (C00544). This reaction is catalyzed by 4-Hydroxyphenylpyruvate dioxygenase (EC 1.13.11.27). The ring structure of homogentisate is subsequently broken and the linear C8 unit degraded in two reaction steps to fumarate and acetoacetate, one citric acid cycle intermediate and one ketone body. Tyrosine transferase (EC 2.6.1.5) is under hormonal control. Glucocorticoids stimulate amino acid degradation in liver which yields fumarate and acetoacetate. Fumarate is converted to oxaloacetate which stimulates gluconeogenesis. Thus liver can synthesize and export glucose into the blood plasma. Acetoacetate is secreted and used by neurons and muscle tissue for oxidative degradation. In enteric bacteria like E.coli, the major degradation product of tyrosine (phenylalanine) is succinate. Also phenol, pyruvate, and free NH3 can be derived by the action of tyrosine-phenol-lyase. Tryptophan Tryptophan degradation in human also yields precursors for glucose synthesis. The first step, catalyzed by tryptophan oxygenase (EC 1.13.11.1), is controlled by cortisol (a glucocorticoid). Tryptophan stimulates its own degradation by allosterically activating tryptophan oxygenase. This is an example of a feed-forward mechanism. The product of the oxygenase reaction is L-Formylkynurenine (C02700), which is further degraded by kynurenine formidase (EC 3.5.1.9). The products are formate and kynurenine (C00328). The latter is degraded to 3-hydroxyanthranilate (C00632) and alanine. This step requires the presence of the cofactor vitamin B6 (pyridoxal). Hydroxyanthranilate can further be decarboxylated to form acetoacetate. Most of hydroxyanthranilate (95%) is used for ketone body formation. The remaining 5 percent go into pyrimidine synthesis (nucleic acid synthesis). Tryptophan is a minor source for the synthesis of NAD (nicotinamide dinucleotide). Most of the nicotinamide is derived from niacin. A daily supply of NAD(P) can be obtained from ~1mg of niacin, while it takes 60mg of the amino acid tryptophan to accomplish the same. Synthesis Amino acid synthesis includes the fixation of nitrogen in form of ammonia and the assimilation of the latter into keto acids to form amino acids by means of glutamate dehydrogenase and glutamine synthetase. The keto acids are provided by glycolysis and citric acid cycle. In total, there are six anabolic pathways for amino acids referred to as biosynthetic families. Many non essential amino acids can directly be obtained by transamination from glutamate to the respective keto acid (e.g. pyruvate to alanine; oxaloacetate to aspartate). Essential amino acids have to be obtained from dietary proteins. Plants and micororganisms have all pathways for the net synthesis of amino acids needed for protein biosynthesis. The synthesis of aromatic ring containing amino acids is discussed below.
Synthesis aromatic amino
acids in E.coli Only microorganisms and plants have the capacity to synthesize aromatic amino acids. Starting from intermediates of glycolysis, the pentose phosphate pathway and/or photosynthesis (calvin cycle), these organisms form the intermediate shikimate (C00493) in several steps. First, shikimate is converted in three steps to chorismate (C00251), which serves as the committed precursor for the synthesis of all three aromatic amino acids, but also for ubiquinone and folate synthesis. The enzyme chorismate mutase (EC 5.4.99.5) catalyzes an intramolecular transferase reaction (isomerization) to form prephenate (C00254), a precursor of tyrosine and phenylalanine. Alternately, chorismate is converted to anthranilate in the presence of glutamine by anthranilate synthase (systematic name: chorismate pyruvate lyase; EC 4.1.3.27). This reaction commits chorismate towards the synthesis of thryptophan. Regulation Amino acid biosynthesis is under allosteric feed back regulation. In general, the end product of a pathway, the amino acid, inhibits the enzyme catalyzing the first (or committed step) of its own biosynthetic pathway. This ensures the energy saving synthesis of building blocks for protein biosynthesis. Accumulating amino acids thus shuts down their biosynthetic activity. A simple feed back mechanism is found in E.coli. Here, isoleucine, which is derived from threonine, inhibits threonine deaminase (EC 4.2.1.16), the committed step in isoleucine biosynthesis which forms alpha-ketobutyrate or oxo-butanoate. For the biosynthetic pathways of more complex and branched aromatic amino acids, however, an analogous sequential feedback mechanism has been elucidated, as found in the bacteria Bacillus subtilis. Simply put, the endpoint of each branch inhibits the first enzymatic step of the immediately preceding branching point. Thus, the aromatic amino acids do not only inhibit their early common pathways leading to shikimate, chorismate, or prephenate intermediates. This assures flexibility for the cell by adjusting the levels of aromatic amino acids according to actual needs. Protein synthesis may need different amounts for phenylalanine than tryptophan. Thus tryptophan may shut down its own biosynthetic branch by inhibiting anthranilate synthase, leaving chorismate mutase unaffected. Chorismate will still be used for phenylalanine or tyrosine formation before it accumulates to shut down the entire pathway. Although E.coli uses the same pathways for the synthesis of aromatic amino acids, it uses a different control mechanism to ensure relative independence between the formation of phenylalanine, tyrosine, and tryptophan. Instead of using this simple sequential feedback mechanism after branching points, E.coli relies on enzyme multiplicity meaning that E.coli uses three different enzymes (2-Dehydro-3-deoxyphosphoheptonate aldolase) for the early synthesis of shikimate. Each enzyme is under allosteric control of its 'own' amino acid end product. Thus the level of enzymes in the cytoplasm determine the level of shikimate and chorismate. For phenylalanine and tyrosine, an additional enzyme multiplicity control is used to allosterically suppress two isoforms of chorismate mutase, one of which is specific for Phe, while the other binds only Tyr. Overall, enzyme multiplicity, too, shows a sequential feedback mechanism because of the multiple branching points of the pathway. Metabolism of sulfur
containing amino acids Two sulfur containing amino acids are used in proteins - methionine and cysteine. Methionine is an essential amino acids for humans. It contributes to the hydrophobicity of a protein. Its sulfur is non reactive. Cysteine in contrast has a highly reactive sulfhydril group which is the cause for disulfide bridges (cysteine pairs) in extracellular proteins (cysteine pairs don't form in the cytoplasm, which has a reducing environment). Cysteines can be part of catalytic sites of proteins or serve as metal ion binding sites, e.g. for Zn coordination in the DNA binding protein domains called Zn-fingers. The metabolism of both sulfur containing amino acids is closely related. Methionine is a precursor for cysteine formation in human liver. Methionine, although it cannot be regenerated from cysteine, can be recycled through methylation of homocysteine (C00155) using a single carbon (methyl-) donor such as betaine (from choline), folate, or vitamin B12 (C05776; Cobalamin (III)) Homocysteine methylation is catalyzed by 5-methyltetrahydrofolate-homocysteine methyltransferase (EC 2.1.1.13) tapping into the one carbon pool metabolism of folate and tetrahydrofolates. Alternately, methionine is regenerated by betaine-homocysteine-S-methyltransferase (EC 2.1.1.5). The methyl donor betaine (C00719) is a derivative of choline. Choline is directly obtained from phosphatidylcholine by the hydrolytic activity of phospholipase C (PLC; EC 3.1.4.3). This reaction pathway is linked to glycine and serine metabolism because the dimethylglycine product is reverted to glycine. The direct recovery of methionine in liver is part of the activated methyl cycle transferring activated methyl groups from a THF donor to S-adenosylmethionine (C00019). This major source for methyl compounds, in form of activated S-adenosylmethionine, comes from dietary methionine. Methionine is linked with ATP to form S-adenosylmethionine releasing both inorganic phosphate and pyrophosphate. This reaction is catalyzed by the enzyme methionine adenosyltransferase EC 2.5.1.6. ATP + L-Methionine + H2O = Orthophosphate + Pyrophosphate + S-Adenosyl-L-methionine The phosphoester hydrolysis fuels the reaction forming an activated methyl group that can be donated to diverse acceptor molecules such as nor-epinephrine (see Neurotransmitters) and ethanolamine (see Phospholipids) to form epinephrine (adrenaline) and choline, respectively. The methyl group is transferred mostly to amino or hydroxyl groups on the acceptor. The amino acid homocysteine cannot be used for protein synthesis and has to be shortened by one carbon unit to cysteine in a process known as trans-sulfuration. In this reaction, serine serves as acceptor. The short lived intermediate cystathionine (C02291; homocysteine -S-serine) is split in a hydrolysis reaction to homoserine and cysteine. Both synthetase (4.2.1.22) and hydrolyse (EC 4.4.1.1) are vitamin B6 (pyridoxal) dependent. The reactions are irreversible. In E.coli cysteine is synthesized from serine in two steps. First, serine is acetylated to acetyl-serine by serine acetyltransferase (EC 2.3.1.30). Acetylserine is converted to L-cysteine by the addition of a sulfhydril group. The enzyme cysteine synthase A (O-acetylserine sulfhydrylase A; EC 4.2.99.8) catalyzes this reaction. Glycine
and tetrahydrofolate metabolism Tetrahydrofolate (THF, C00101) as discussed above is an important carrier of activated one-carbon units. While S-adenosylmethionine only transfers methyl groups, THF carries a great variety of activated one-carbon units of different oxidation states:
Carboxylation (-COOH) cannot be mediated by THF, instead always requires biotin as coenzyme. One-carbon units are interconvertibel (can be reduced or oxidized; see one carbon pool metabolism) and are activated on THF on the nitrogens N5 or N10 or both (methylene). The major one-carbon acceptor molecules are found in the biosynthetic
pathways of methionine, glycine, purines, and deoxythymine. The
example for glycine is shown below. Glycine synthase catalyzes
a reversible reaction and thus produces either glycine or one carbon
units and ammonium. |
