Formation of glycine and aminoacetone from L-threonine by rat liver mitochondria

Threonine is a precursor of glycine in the rat, but the metabolic pathway involved is unclear. To elucidate this pathway, the biosynthesis of glycine, and of aminoacetone, from L-threonine were studied in rat liver mitochondrial preparations of differing integrities. In the absence of added cofactors, intact mitochondria formed glycine and aminoacetone in approximately equal amounts from 20 mM L-threonine, but exogenous NAD+ decreased and CoA increased the ratio of glycine to aminoacetone formed. In intact and freeze-thawed mitochondria, the ratio of glycine to aminoacetone formed was markedly sensitive to the concentration of L-threonine, glycine being the major product at low L-threonine concentrations. Disruption of mitochondrial integrity by sonication (1 min) decreased the ratio of glycine to aminoacetone formed, and in 20000 X g supernatant fractions from sonicated (3 min) mitochondria, aminoacetone was the major product. The main non-nitogenous two-carbon compound detected when intact mitochondria catabolized L-threonine to glycine was acetate, which was probably derived from deacylation of acetyl-CoA. These results suggest that glycine formation from L-threonine in rat liver mitochondria occurred primarily by the coupled activities of threonine dehydrogenase and 2-amino-3-oxobutyrate CoA-ligase, the extent of coupling between the enzymes being dependent upon a close physical relationship and upon the flux through the dehydrogenase reaction. In vivo glycine synthesis would predominate, and aminoacetone would be a minor product.     

Purification and properties of aminoacetone synthetase from beef liver mitochondria

Aminoacetone synthetase from beef liver mitochondria was purified to homogeneity and shown to be a member of the pyridoxal 5'-phosphate-dependent family of enzymes. This enzyme catalyzes the condensation of glycine and acetyl-CoA to produce CO2, CoA, and the stable product aminoacetone. Bovine aminoacetone synthetase is a dimer (Mr 56,000) of identical subunits and contains 2 mol of pyridoxal phosphate/mol of dimer. The holoenzyme was resolved by dialysis against cysteine and has a pI of 5.2. The holoenzyme shows an absorption maximum at 428 nm which undergoes a shift to 335 nm when reduced with sodium borohydride. The Km values of glycine and acetyl-CoA were 22 mM and 53 microM, respectively. Initial velocity studies indicate that the condensation reaction proceeds by an ordered mechanism. With the exception of aminomalonate, bovine aminoacetone synthetase acts specifically on glycine and acetyl-CoA. Coupled reactions of purified bovine aminoacetone synthetase and porcine L-threonine dehydrogenase demonstrated the interconversion of threonine and glycine.     

Growth, enzyme levels, and some metabolic properties of an Escherichia coli mutant grown on L-threonine as the sole carbon source.

A mutant of Escherichia coli (designated E. coli SBD-76) that utilizes L-threonine as the sole carbon source was isolated. In contrast with levels in extracts of wild-type cells, the levels of threonine dehydrogenase in extracts of this mutant were 100-fold higher than levels of threonine aldolase or degradative threonine dehydratase. Catabolite repression of threonine dehydrogenase was manifested in wild-type, but not SBD-76, cells. For purposes of isolating enzymes, large quantities of SBD-76 cells with the elevated threonine dehydrogenase level could be grown in a fermentor in modified Fraser medium containing 1% glycerol, rather than in the 0.2% L-threonine minimal medium used to isolate the mutant. SBD-76 cells grown on L-threonine excreted glycine and aminoacetone into the medium, and extracts of the mutant strain catalyzed a quantitative conversion of L-threonine to glycine and aminoacetone.     

Threonine formation via the coupled activity of 2-amino-3-ketobutyrate coenzyme A lyase and threonine dehydrogenase.

The enzymes L-threonine dehydrogenase and 2-amino-3-ketobutyrate coenzyme A (CoA) lyase are known to catalyze the net conversion of L-threonine plus NAD+ plus CoA to NADH plus glycine plus acetyl-CoA. When homogeneous preparations of these two enzymes from Escherichia coli were incubated together for 40 min at 25 degrees C with glycine, acetyl-CoA, and NADH, a 36% decrease in the level of glycine (with concomitant NADH oxidation) was matched by formation of an equivalent amount of threonine, indicating that this coupled sequence of enzyme-catalyzed reactions is reversible in vitro. Several experimental factors that affect the efficiency of this conversion in vitro were examined. A constructed strain of E. coli, MD901 (glyA thrB/C tdh), was unable to grow unless both glycine and threonine were added to defined rich medium. Introduction of the plasmid pDR121 (tdh+kbl+) into this strain enabled the cells to grow in the presence of either added glycine or threonine, indicating that interconversion of these two amino acids occurred. Threonine that was isolated from the total pool of cellular protein of MD901/pDR121 had the same specific radioactivity as the [14C]glycine added to the medium, establishing that threonine was formed exclusively from glycine in this strain. Comparative growth rate studies with several strains of E. coli containing plasmid pDR121, together with the finding that kcat values of pure E. coli 2-amino-3-ketobutyrate CoA lyase favor the cleavage of 2-amino-3-ketobutyrate over its formation by a factor of 50, indicate that the biosynthesis of threonine is less efficient than glycine formation via the coupled threonine dehydrogenase-2-amino-3-ketobutyrate lyase reactions.     

Formation of glycine and aminoacetone from l-threonine by rat liver mitochondria

Threonine is a precursor of glycine in the rat, but the metabolic pathway involved is unclear. To elucidate this pathway, the biosynthesis of glycine, and of aminoacetone, from l-threonine were studied in rat liver mitochondrial preparations of differing integrities. In the absence of added cofactors, intact mitochondria formed glycine and aminoacetone in approximately equal amounts from 20 mM l-threonine, but exogenous NAD⁺ decreased and CoA increased the ratio of glycine to aminoacetone formed. In intact and freeze-thawed mitochondria, the ratio of glycine to aminoacetone formed was markedly sensitive to the concentration of l-threonine, glycine being the major product at low l-threonine concentrations. Disruption of mitochondrial integrity by sonication (1 min) decreased the ratio of glycine to aminoacetone formed, and in 20 000 × g supernatant fractions from sonicated (3 min) mitochondria, aminoacetone was the major product. The main non-nitogenous tow-carbon compound detected when intact mitochondria catabolized l-threonine to glycine was acetate, which was probably derived from deacylation of acetyl-CoA. These results suggest that glycine formation from l-threonine in rat liver mitochondria occured primarily by the coupled activities of threonine dehydrogenase and 2-amino-3-oxobutyrate CoA-ligase, the extent of coupling between the enzymes being dependent upon a close physical relationship and upon the flux through the dehydrogenase reaction. In vivo glycine synthesis would predominate, and aminoacetone would be a minor product.     

L-Threonine dehydrogenase from goat liver. Feedback inhibition by methylglyoxal

L-Threonine dehydrogenase, which forms aminoacetone from L-threonine and NAD, has been extensively purified from goat liver. A feedback inhibition of this enzyme has been observed with methylglyoxal. Kinetic data and other experiments indicate that methylglyoxal acts at a site other than the active site of the enzyme. The enzyme contains a single subunit of Mr 89,000. The apparent Km values of the enzyme for L-threonine and NAD were found to be 5.5 and 1 mM, respectively.     

Metabolism of 2-oxoaldehydes in yeasts. Possible role of glycolytic bypath as a detoxification system in L-threonine catabolism by Saccharomyces cerevisiae

L-Threonine catabolism by Saccharomyces cerevisiae was studied to determine the role of glycolytic bypath as a detoxyfication system of 2-oxoaldehyde (methylglyoxal) formed from L-threonine catabolism. During the growth on L-threonine as a sole source of nitrogen, a large amount of aminoacetone was accumulated in the culture. The enzymatic analyses indicated that L-threonine was converted into either acetaldehyde and glycine by threonine aldolase or 2-aminoacetoacetate by NAD-dependent threonine dehydrogenase. Glycine formed was condensed with acetyl-CoA by aminoacetone synthase to form 2-aminoacetoacetate, a labile compound spontaneously decarboxylated into aminoacetone. The enzyme activities of the glycolytic bypath of the cells grown on L-threonine were considerably higher than those of the cells grown on ammonium sulfate as a nitrogen source. The result indicated the possible role of glycolytic bypath as a detoxification system of methylglyoxal formed from L-threonine catabolism.     

Utilization of l-threonine by a species of Arthrobacter. A novel catabolic role for `aminoacetone synthase''

1. A species of Arthrobacter (designated Arthrobacter 9759) was isolated from soil by its ability to grow aerobically on l-threonine as sole source of carbon atoms, nitrogen atoms and energy; the organism also grew well on other sources of carbon atoms including glycine, but no growth was obtainable on aminoacetone or dl-1-aminopropan-2-ol. 2. During growth on threonine, (14)C from l-[U-(14)C]threonine was rapidly incorporated into glycine and citrate, and thereafter into serine, alanine, aspartate and glutamate. 3. With extracts of threonine-grown cells supplied with l-[U-(14)C]threonine, evidence was obtained of the NAD and CoA-dependent catabolism of l-threonine to produce acetyl-CoA plus glycine. Short-term incorporation studies in which [2-(14)C]acetate and [2-(14)C]glycine were supplied (a) to cultures growing on threonine, and (b) to extracts of threonine-grown cells, showed that the acetyl-CoA was metabolized via the tricarboxylic acid cycle and glyoxylate cycle whereas the glycine was converted into pyruvate via the folate-dependent ;serine pathway'. 4. The threonine-grown organism contained ;biosynthetic' threonine dehydratase and a potent NAD-linked l-threonine dehydrogenase but possessed no l-threonine aldolase activity. 5. Evidence was obtained that the acetyl-CoA and glycine produced from l-threonine had their immediate origin in the alpha-amino-beta-oxobutyrate formed by the threonine dehydrogenase; the CoA-dependent cleavage of this compound was catalysed by an alpha-amino-beta-oxobutyrate CoA-ligase, which was identified with ;aminoacetone synthase'. A continuous spectrophotometric assay of this enzyme was developed, and it was found to be inducibly synthesized only during growth on threonine and not during growth on acetate plus glycine. 6. By using a reconstituted mixture of separately purified l-threonine dehydrogenase and alpha-amino-beta-oxobutyrate CoA-ligase (i.e. ;aminoacetone synthase'), l-[U-(14)C]threonine was broken down to [(14)C]glycine plus [(14)C]acetyl-CoA (trapped as [(14)C]citrate). 7. There was no evidence of aminoacetone metabolism by Arthrobacter 9759 even though a small amount of this amino ketone appeared in the culture medium during growth on threonine.     

Threonine degradation by Serratia marcescens.

The wild strain of Serratia marcescens rapidly degraded threonine and formed aminoacetone in a medium containing glucose and urea. Extracts of this strain showed high threonine dehydrogenase and "biosynthetic" threonine deaminase activities, but no threonine aldolase activity. Threonine dehydrogenase-deficient strain Mu-910 was selected among mutants unable to grow on threonine as the carbon source. This strain did not form aminoacetone from threonine, but it slowly degraded threonine. Strain D-60, deficient in both threonine dehydrogenase and threonine deaminase, was derived from strain Mu-910 and barely degraded threonine. A glycine-requiring strain derived from the wild strain grew in minimal medium containing threonine as the glycine source, whereas a glycine-requiring strain derived from strain Mu-910 did not grow. This indicates that threonine dehydrogenase participates in glycine formation from threonine (via alpha-amino-beta-ketobutyrate) as well as in threonine degradation to aminoacetone.     

Oxidative DNA damage induced by aminoacetone, an amino acid metabolite

We investigated DNA damage induced by aminoacetone, a metabolite of threonine and glycine. Pulsed-field gel electrophoresis revealed that aminoacetone caused cellular DNA cleavage. Aminoacetone increased the amount of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) in human cultured cells in a dose-dependent manner. The formation of 8-oxodG in calf thymus DNA increased due to aminoacetone only in the presence of Cu(II). DNA ladder formation was observed at higher concentrations of aminoacetone than those causing DNA cleavage. Flow cytometry showed that aminoacetone enhanced the generation of hydrogen peroxide (H2O2) in cultured cells. Aminoacetone caused damage to 32P-5'-end-labeled DNA fragments, obtained from the human c-Ha-ras-1 and p53 genes, at cytosine and thymine residues in the presence of Cu(II). Catalase and bathocuproine inhibited DNA damage, suggesting that H2O2 and Cu(I) were involved. Analysis of the products generated from aminoacetone revealed that aminoacetone underwent Cu(II)-mediated autoxidation in two different pathways: the major pathway in which methylglyoxal and NH+4 are generated and the minor pathway in which 2,5-dimethylpyrazine is formed through condensation of two molecules of aminoacetone. These findings suggest that H2O2 generated by the autoxidation of aminoacetone reacts with Cu(I) to form reactive species capable of causing oxidative DNA damage.     

Bacterial catabolism of threonine. Threonine degradation initiated by L-threonine-NAD+ oxidoreductase.

1. Isolates representing seven bacterial genera capable of growth on L-threonine medium, and possessing high L-threonine 3-dehydrogenase activity, were examined to elucidate the catabolic route. 2. The results of growth, manometric and enzymic experiments indicated the catabolism of L-threonine by cleavage to acetyl-CoA plus glycine, the glycine being further metabolized via L-serine to pyruvate, in all cases. No evidence was obtained of a role for aminoacetone in threonine catabolism or for the metabolism of glycine by the glycerate pathway. 3. The properties of a number of key enzymes in L-threonine catabolism were investigated. The inducibly formed L-threonine 3-dehydrogenase, purified from Corynebacterium sp. B6 to a specific activity of about 30-35 mumol of product formed/min per mg of protein, exhibited a sigmoid kinetic response to substrate concentration. The half-saturating concentration of substrate, [S]0.5, was 20mM and the Hill constant (h) was 1.50. The Km for NAD+ was 0.8mM. The properties of the enzyme were studied in cell-free extracts of other bacteria. 4. New assays for 2-amino-3-oxobutyrate-CoA ligase were devised. The Km for CoA was determined for the first time and found to be 0.14mM at pH8, for the enzyme from Corynebacterium sp. B6. Evidence was obtained for the efficient linkage of the dehydrogenase and ligase enzymes. Cell-free extracts all possessed high activities of the inducibly formed ligase. 5. L-Serine hydroxymethyltransferase was formed constitutively by all isolates, whereas formation of the 'glycine-cleavage system' was generally induced by growth on L-threonine or glycine. The coenzyme requirements of both enzymes were established, and their linked activity in the production of L-serine from glycine was demonstrated by using extracts of Corynebacterium sp. B6. 6. L-Serine dehydratase, purified from Corynebacterium sp. B6 to a specific activity of about 4mumol of product formed/min per mg of protein, was found to exhibit sigmoid kinetics with an [S]0.5 of about 20mM and h identical to 1.4. Similar results were obtained with enzyme preparations from all isolates. The enzyme required Mg2+ for maximum activity, was different from the L-threonine dehydratase also detectable in extracts, and was induced by growth on L-threonine or glycine.     

Catabolism of threonine in mammals by coupling of l-threonine 3-dehydrogenase with 2-amino-3-oxobutyrate-CoA ligase

There is doubt about the l-threonine 3-dehydrogenase (EC 1.1.1.103) and threonine aldolase (EC 2.1.2.1) catabolic pathways of l-threonine in mammals which are believed to produce aminoacetone and glycine plus acetaldehyde, respectively. l-Threonine 3-dehydrogenase in disrupted guinea-pig liver mitochondria was investigated in a reaction mixture containing l-threonine without and with CoA and oxaloacetate; l-[U-¹⁴C]threonine was included in four similar experiments for autoradiograms. Threonine aldolase was examined in similar mitochondria from liver and kidney. CoA reduced the aminoacetone formed from l-threonine to 10–14% and CoA plus oxaloacetate produced citrate (from CoASAc) in approximately equal amounts to the decrease in aminoacetone. Autoradiograms confirmed the decrease in aminoacetone with the simultaneous appearance of citrate and glycine. No evidence was obtained that threonine aldolase catabolised l-threonine at the concentration used to assay the dehydrogenase. It is concluded that 2-amino-3-oxobutyrate (precursor of aminoacetone), which is produced from l-threonine by l-threonine 3-dehydrogenase, undergoes CoA-dependent cleavage to glycine and CoASAc by 2-amino-3-oxobutyrate-CoA ligase. The results suggest that the coupling of these enzymes provides a new pathway for the catabolism of threonine in mammals.     

The human L-threonine 3-dehydrogenase gene is an expressed pseudogene

Background  

L-threonine is an indispensable amino acid. One of the major L-threonine degradation pathways is the conversion of L-threonine via 2-amino-3-ketobutyrate to glycine. L-threonine dehydrogenase (EC 1.1.1.103) is the first enzyme in the pathway and catalyses the reaction: L-threonine + NAD⁺ = 2-amino-3-ketobutyrate + NADH. The murine and porcine L-threonine dehydrogenase genes (TDH) have been identified previously, but the human gene has not been identified.     

The sulfhydryl content of L-threonine dehydrogenase from Escherichia coli K-12: relation to catalytic activity and Mn2+ activation

When oxidized to cysteic acid by performic acid or converted to carboxymethylcysteine by alkylation of the reduced enzyme with iodoacetate, a total of six half-cystine residues/subunit are found in L-threonine dehydrogenase (L-threonine: NAD+ oxidoreductase, EC 1.1.1.103; L-threonine + NAD(+)----2-amino-3-oxobutyrate + NADH) from Escherichia coli K-12. Of this total, two exist in disulfide linkage, whereas four are titratable under denaturing conditions by dithiodipyridine, 5,5'-dithiobis(2-nitrobenzoic acid), or p-mercuribenzoate. The kinetics of enzyme inactivation and of modification by the latter two reagents indicate that threonine dehydrogenase has no free thiols that selectively react with bulky compounds. While incubation of the enzyme with a large excess of iodoacetamide causes less than 10% loss of activity, the native dehydrogenase is uniquely reactive with and completely inactivated by iodoacetate. The rate of carboxymethylation by iodoacetate of one -SH group/subunit is identical with the rate of inactivation and the carboxymethylated enzyme is no longer able to bind Mn2+. NADH (0.5 mM) provides 40% protection against this inactivation; 60 to 70% protection is seen in the presence of saturating levels of NADH plus L-threonine. Such results coupled with an analysis of the kinetics of inactivation caused by iodoacetate are interpreted as indicating the inhibitor first forms a reversible complex with a positively charged moiety in or near the microenvironment of a reactive -SH group in the enzyme before irreversible alkylation occurs. Specific alkylation of one -SH group/enzyme subunit apparently causes protein conformational changes that entail a loss of catalytic activity and the ability to bind Mn2+.     

Catabolism of threonine in mammals by coupling of l-threonine 3-dehydrogenase with 2-amino-3-oxobutyrate-CoA ligase

There is doubt about the l-threonine 3-dehydrogenase (EC 1.1.1.103) and threonine aldolase (EC 2.1.2.1) catabolic pathways of l-threonine in mammals which are believed to produce aminoacetone and glycine plus acetaldehyde, respectively. l-Threonine 3-dehydrogenase in disrupted guinea-pig liver mitochondria was investigated in a reaction mixture containing l-threonine without and with CoA and oxaloacetate; l-[U-¹⁴C]threonine was included in four similar experiments for autoradiograms. Threonine aldolase was examined in similar mitochondria from liver and kidney. CoA reduced the aminoacetone formed from l-threonine to 10–14% and CoA plus oxaloacetate produced citrate (from CoASAc) in approximately equal amounts to the decrease in aminoacetone. Autoradiograms confirmed the decrease in aminoacetone with the simultaneous appearance of citrate and glycine. No evidence was obtained that threonine aldolase catabolised l-threonine at the concentration used to assay the dehydrogenase. It is concluded that 2-amino-3-oxobutyrate (precursor of aminoacetone), which is produced from l-threonine by l-threonine 3-dehydrogenase, undergoes CoA-dependent cleavage to glycine and CoASAc by 2-amino-3-oxobutyrate-CoA ligase. The results suggest that the coupling of these enzymes provides a new pathway for the catabolism of threonine in mammals.     

L-threonine dehydrogenase from Escherichia coli K-12: thiol-dependent activation by Mn2+

Addition of 1 mM Mn2+ to all solutions in the final chromatographic step used to purify L-threonine dehydrogenase (L-threonine:NAD+ oxidoreductase, EC 1.1.1.103) from extracts of Escherichia coli K-12 routinely provides 30-40 mg of pure enzyme per 100 g wet weight of cells with specific activity = 20-30 units/mg. Enzyme dialyzed exhaustively against buffers containing Chelex-100 resin has a specific activity = 8 units/mg and contains 0.003 or 0.02 mol of Mn2+/mol of enzyme as determined by radiolabeling studies with 54Mn2+ or by atomic absorption spectroscopy, respectively. Dehydrogenase activity is completely abolished by low concentrations of either Hg2+ or Ag+; of a large spectrum of other metal ions tested, only Mn2+ and Cd2+ have an activating effect. Activation of threonine dehydrogenase by Mn2+ is thiol-dependent and is saturable with an activation Kd = 9.0 microM and a Vmax = 105 units/mg. Stoichiometry of Mn2+ binding was found to be 0.86 mol of Mn2+/mol of enzyme subunit with a dissociation constant (Kd) = 8.5 microM. Mn2+ appears to interact directly with threonine dehydrogenase; gel filtration studies with the dehydrogenase plus 54Mn2+ in the presence of either NAD+, NADH, L-threonine, or combinations thereof show that only Mn2+ coelutes with the enzyme whereas all other ligands elute in the salt front and the stoichiometry of the dehydrogenase-Mn2+ interaction is not affected in any instance. A theoretical curve fit to data for the pH-activity profile of Mn2+-saturated enzyme has a pKa = 7.95 for one proton ionization. The data establish L-threonine dehydrogenase of E. coli to be a metal ion activated enzyme.     

The oxidation of aminoacetone by a species of Arthrobacter

1. A micro-organism similar to Arthrobacter globiformis has been isolated from sewage by elective growth on a medium containing l-threonine as sole source of carbon and nitrogen. 2. Washed cell suspensions of the organism catalyse the complete disappearance of aminoacetone from the medium and its almost complete oxidation. 3. In the presence of iodoacetate, aminoacetone disappearance is accompanied by the accumulation of methylglyoxal, about 70% of the aminoacetone removed being accounted for in this way. 4. It is suggested that the conversion of aminoacetone into methylglyoxal is catalysed by an amine oxidase.     

Inactivation of Escherichia coli L-threonine dehydrogenase by 2,3-butanedione. Evidence for a catalytically essential arginine residue

Incubation of homogeneous preparations of L-threonine dehydrogenase from Escherichia coli with 2,3-butanedione, 2,3-pentanedione, phenylglyoxal, or 1,2-cyclohexanedione causes a time- and concentration-dependent loss of enzymatic activity; plots of log percent activity remaining versus time are linear to greater than 90% inactivation, indicative of pseudo-first order inactivation kinetics. The reaction order with respect to the concentration of modifying reagent is approximately 1.0 in each case suggesting that the loss of catalytic activity is due to one molecule of modifier reacting with each active unit of enzyme. Controls establish that this inactivation is not due to modifier-induced dissociation or photoinduced nonspecific alteration of the dehydrogenase. Essentially the same Km but decreased Vmax values are obtained when partially inactivated enzyme is compared with native. NADH (25 mM) and NAD+ (70 mM) give full protection against inactivation whereas much higher concentrations (i.e. 150 mM) of L-threonine or L-threonine amide provide a maximum of 80-85% protection. Amino acid analyses coupled with quantitative sulfhydryl group determinations show that enzyme inactivated 95% by 2,3-butanedione loses 7.5 arginine residues (out of 16 total)/enzyme subunit with no significant change in other amino acid residues. In contrast, only 2.4 arginine residues/subunit are modified in the presence of 80 mM NAD+. Analysis of the course of modification and inactivation by the statistical method of Tsou (Tsou, C.-L. (1962) Sci. Sin. 11, 1535-1558) demonstrates that inactivation of threonine dehydrogenase correlates with the loss of 1 "essential" arginine residue/subunit which quite likely is located in the NAD+/NADH binding site.     

Inhibition and regulation of rat liver L-threonine dehydrogenase by different fatty acids and their derivatives.

Rat liver L-threonine dehydrogenase is a mitochondrial enzyme which transforms L-threonine either into aminoacetone or into acetyl-CoA. We show that it is inhibited by several fatty acids and their derivatives: short chain fatty acids, L-2-hydroxybutyrate and D-3-hydroxybutyrate, long chain fatty acids, such as lauric acid, myristic acid, palmitic and stearic acids, bicarboxylic acids such as malonic acid and its derivatives methyl- and hydroxymalonic acids. The inhibition occurs at low and physiological concentrations of such compounds, which are normally present and metabolized in mitochondria. It presumably plays a role in the physiology of acetyl-CoA-dependent formation of fatty acids and ketobodies, in L-threonine-dependent gluconeogenesis, and in the regulation of L-threonine metabolism by L-threonine dehydrogenase and L-threonine deaminase.