Amino Acid Catabolic Pathways Research Articles

Factors influencing cell fatty acid composition and A40926 antibiotic complex production in Nonomuraea sp. ATCC 39727

A40926 is a glycopeptide antibiotic complex consisting of several structurally related factors. It is produced by fermentation of Nonomuraea sp. ATCC 39727 and the complex components differ in the structure of the fatty acid moiety linked to the aminoglucuronic acid unit. In previous work, we observed that the production of single factors in glycopeptide antibiotic complexes could be selectively enhanced by the addition of suitable precursors to the culture medium. In this contribution, we examine the effects of branched amino acid addition to fermentation of Nonomuraea sp. in a chemically defined minimal medium. The changes in the composition of cell fatty acids correlate to the fatty acid distribution within the A40926 complex in diverse cultivation conditions. Nonomuraea sp. prefers isobutyric, butyric and propionic acids as initiators of fatty acid biosynthesis. The relative amount of the produced fatty acids is significantly influenced by the availability of intermediates or final products from the amino acid catabolic pathways. Antibiotic complex composition closely reflects the cell fatty acid pattern, in agreement with the assumption that the antibiotic fatty acid moieties are synthesized by shortening the chain of cell fatty acids.

Analysis of Growth of Lactobacillus plantarum WCFS1 on a Complex Medium Using a Genome-scale Metabolic Model

A genome-scale metabolic model of the lactic acid bacterium Lactobacillus plantarum WCFS1 was constructed based on genomic content and experimental data. The complete model includes 721 genes, 643 reactions, and 531 metabolites. Different stoichiometric modeling techniques were used for interpretation of complex fermentation data, as L. plantarum is adapted to nutrient-rich environments and only grows in media supplemented with vitamins and amino acids. (i) Based on experimental input and output fluxes, maximal ATP production was estimated and related to growth rate. (ii) Optimization of ATP production further identified amino acid catabolic pathways that were not previously associated with free-energy metabolism. (iii) Genome-scale elementary flux mode analysis identified 28 potential futile cycles. (iv) Flux variability analysis supplemented the elementary mode analysis in identifying parallel pathways, e.g. pathways with identical end products but different co-factor usage. Strongly increased flexibility in the metabolic network was observed when strict coupling between catabolic ATP production and anabolic consumption was relaxed. These results illustrate how a genome-scale metabolic model and associated constraint-based modeling techniques can be used to analyze the physiology of growth on a complex medium rather than a minimal salts medium. However, optimization of biomass formation using the Flux Balance Analysis approach, reported to successfully predict growth rate and by product formation in Escherichia coli and Saccharomyces cerevisiae, predicted too high biomass yields that were incompatible with the observed lactate production. The reason is that this approach assumes optimal efficiency of substrate to biomass conversion, and can therefore not predict the metabolically inefficient lactate formation.

A branched-chain amino acid aminotransferase gene isolated from Hordeum vulgare is differentially regulated by drought stress

Differential display was used to isolate cDNA clones showing differential expression in response to ABA, drought and cold in barley seedling shoots. One drought-regulated cDNA clone (DD12) was further analyzed and found to encode a branched-chain amino acid aminotransferase (HvBCAT-1). A genomic clone was isolated by probing the Morex BAC library with the cDNA clone DD12 and the structure of Hvbcat-1 was elucidated. The coding region is interrupted by six introns and contains a predicted mitochondrial transit peptide. Hvbcat1 was mapped to chromosome 4H. A comparison was made to rice and Arabidopsis genes to identify conserved structural patterns. Complementation of a yeast (Saccharomyces cerevisiae) double knockout strain revealed that HvBCAT-1 can function as the mitochondrial (catabolic) BCATs in vivo. Transcript levels of Hvbcat-1, increased in response to drought stress. As the first enzyme in the branched-chain amino acid (BCAA) catabolic pathway, HvBCAT-1 might have a role in the degradation of BCAA. Degradation of BCAA could serve as a detoxification mechanism that maintains the pool of free branched-chain amino acids at low and non toxic levels, under drought stress conditions.

Identification of the Leucine-to-2-Methylbutyric Acid Catabolic Pathway of Lactococcus lactis

Nutrient starvation and nonculturability in bacteria lead to changes in metabolism not found during the logarithmic phase. Substrates alternate to those used during growth are metabolized in these physiological states, yielding secondary metabolites. In firmicutes and actinobacteria, amino acid catabolic pathways are induced during starvation and nonculturability. Examination of lactococci showed that the population entered a nonculturable state after carbohydrate depletion and was incapable of growth on solid media; however, the cells gained the ability to produce branched-chain fatty acids from amino acids. Gene expression profiling and in silico pathway analysis coupled with nuclear magnetic resonance spectroscopy were used to delineate the leucine catabolic pathway. Lactococci produced acetic and propionic acid during logarithmic growth and starvation. At the onset of nonculturability, 2-methylbutyric acid was produced via hydroxymethyl-glutaryl-coenzyme A (CoA) and acetyl-CoA, along with ATP and oxidation/reduction precursors. Gene expression profiling and genome sequence analysis showed that lactococci contained redundant genes for branched-chain fatty acid production that were regulated by an unknown mechanism linked to carbon metabolism. This work demonstrated the ability of a firmicute to induce new metabolic capabilities in the nonculturable state for producing energy and intermediates needed for transcription and translation. Phylogenetic analyses showed that homologues of these enzymes and their functional motifs were widespread across the domains of life.

Amino Acid Catabolic Pathways of Lactic Acid Bacteria

Lactic acid bacteria (LAB) constitute a diverse group of Gram positive obligately fermentative microorganisms which include both beneficial and pathogenic strains. LAB generally have complex nutritional requirements and therefore they are usually associated with nutrient-rich environments such as animal bodies, plants and foodstuffs. Amino acids represent an important resource for LAB and their utilization serves a number of physiological roles such as intracellular pH control, generation of metabolic energy or redox power, and resistance to stress. As a consequence, the regulation of amino acid catabolism involves a wide set of both general and specific regulators and shows significant differences among LAB. Moreover, due to their fermentative metabolism, LAB amino acid catabolic pathways in some cases differ significantly from those described in best studied prokaryotic model organisms such as Escherichia coli or Bacillus subtilis. Thus, LAB amino acid catabolism constitutes an interesting case for the study of metabolic pathways. Furthermore, LAB are involved in the production of a great variety of fermented products so that the products of amino acid catabolism are also relevant for the safety and the quality of fermented products.

Methionine catabolism inSaccharomyces cerevisiae

The catabolism of methionine to methionol and methanethiol in Saccharomyces cerevisiae was studied using (13)C NMR spectroscopy, GC-MS, enzyme assays and a number of mutants. Methionine is first transaminated to alpha-keto-gamma-(methylthio)butyrate. Methionol is formed by a decarboxylation reaction, which yields methional, followed by reduction. The decarboxylation is effected specifically by Ydr380wp. Methanethiol is formed from both methionine and alpha-keto-gamma-(methylthio)butyrate by a demethiolase activity. In all except one strain examined, demethiolase was induced by the presence of methionine in the growth medium. This pathway results in the production of alpha-ketobutyrate, a carbon skeleton, which can be re-utilized. Hence, methionine catabolism is more complex and economical than the other amino acid catabolic pathways in yeast, which use the Ehrlich pathway and result solely in the formation of a fusel alcohol.

Ability of thermophilic lactic acid bacteria to produce aroma compounds from amino acids.

Although a large number of key odorants of Swiss-type cheese result from amino acid catabolism, the amino acid catabolic pathways in the bacteria present in these cheeses are not well known. In this study, we compared the in vitro abilities of Lactobacillus delbrueckii subsp. lactis, Lactobacillus helveticus, and Streptococcus thermophilus to produce aroma compounds from three amino acids, leucine, phenylalanine, and methionine, under mid-pH conditions of cheese ripening (pH 5.5), and we investigated the catabolic pathways used by these bacteria. In the three lactic acid bacterial species, amino acid catabolism was initiated by a transamination step, which requires the presence of an alpha-keto acid such as alpha-ketoglutarate (alpha-KG) as the amino group acceptor, and produced alpha-keto acids. Only S. thermophilus exhibited glutamate dehydrogenase activity, which produces alpha-KG from glutamate, and consequently only S. thermophilus was capable of catabolizing amino acids in the reaction medium without alpha-KG addition. In the presence of alpha-KG, lactobacilli produced much more varied aroma compounds such as acids, aldehydes, and alcohols than S. thermophilus, which mainly produced alpha-keto acids and a small amount of hydroxy acids and acids. L. helveticus mainly produced acids from phenylalanine and leucine, while L. delbrueckii subsp. lactis produced larger amounts of alcohols and/or aldehydes. Formation of aldehydes, alcohols, and acids from alpha-keto acids by L. delbrueckii subsp. lactis mainly results from the action of an alpha-keto acid decarboxylase, which produces aldehydes that are then oxidized or reduced to acids or alcohols. In contrast, the enzyme involved in the alpha-keto acid conversion to acids in L. helveticus and S. thermophilus is an alpha-keto acid dehydrogenase that produces acyl coenzymes A.

Effects of liver failure on branched-chain α-keto acid dehydrogenase complex in rat liver and muscle: comparison between acute and chronic liver failure

Background/Aims: Branched-chain α-keto acid dehydrogenase (BCKDH) complex catalyses the committed step in the branched-chain amino acid (BCAA) catabolic pathway. In many cases of liver failure, the serum BCAAs/aromatic amino acids ratio (Fisher's ratio) decreases, and BCAAs have been administered to patients with liver failure to correct this ratio. We conducted an animal study to examine whether the effects on hepatic BCKDH complex differ between acute liver failure (ALF) and chronic liver failure (CLF). Methods: ALF and CLF was induced in rats by a single high-dose injection and 21 weeks of repeated low-dose injections of carbon tetrachloride, respectively. Plasma BCAA and branched-chain α-keto acid (BCKA) levels, and activities and protein amounts of hepatic BCKDH complex and kinase were measured. Results: ALF was characterized by elevated plasma BCAA and BCKA levels and decreased hepatic BCKDH activity. CLF was characterized by decreased plasma BCAA and BCKA levels and increased hepatic BCKDH activity. This increase in BCKDH activity in CLF was associated with the decreased BCKDH kinase, which is responsible for the BCKDH inactivation. Conclusions: The results obtained in the present study suggest that BCAA catabolism is suppressed in ALF and increased in CLF.

Effects of liver failure on the enzymes in the branched-chain amino acid catabolic pathway

Branched-chain α-keto acid dehydrogenase (BCKDH) complex catalyzes the committed step of the catabolism of branched-chain amino acids (BCAA). The liver cirrhosis chemically induced in rats raised the activity of hepatic BCKDH complex and decreased plasma BCAA and branched-chain α-keto acid concentrations, suggesting that the BCAA requirement is increased in liver cirrhosis. Since the effects of liver cirrhosis on the BCKDH complex in human liver are different from those in rat liver, further studies are needed to clarify the differences between rats and humans. In the valine catabolic pathway, crotonase and β-hydroxyisobutyryl-CoA hydrolase are very important to regulate the toxic concentration of mitochondrial methacrylyl-CoA, which occurs in the middle part of valine pathway and highly reacts with free thiol compounds. Both enzyme activities in human and rat livers are very high compared to that of BCKDH complex. It has been found that both enzyme activities in human livers were significantly reduced by liver cirrhosis and hepatocellular carcinoma, suggesting a decrease in the capability to dispose methacrylyl-CoA. The findings described here suggest that alterations in hepatic enzyme activities in the BCAA catabolism are associated with liver failure.

Branched-chain amino acid catabolism: unique segregation of pathway enzymes in organ systems and peripheral nerves.

We have examined the localization of the first two enzymes in the branched-chain amino acid (BCAA) catabolic pathway: the branched-chain aminotransferase (BCAT) isozymes (mitochondrial BCATm and cytosolic BCATc) and the branched-chain alpha-keto acid dehydrogenase (BCKD) enzyme complex. Antibodies specific for BCATm or BCATc were used to immunolocalize the respective isozymes in cryosections of rat tissues. BCATm was expressed in secretory epithelia throughout the digestive tract, with the most intense expression in the stomach. BCATm was also strongly expressed in secretory cells of the exocrine pancreas, uterus, and testis, as well as in the transporting epithelium of convoluted tubules in kidney. In muscle, BCATm was located in myofibrils. Liver, as predicted, was not immunoreactive for BCATm. Unexpectedly, BCATc was localized in elements of the autonomic innervation of the digestive tract, as well as in axons in the sciatic nerve. The distributions of BCATc and BCATm did not overlap. BCATm-expressing cells also expressed the second enzyme of the BCAA catabolic pathway, BCKD. In selected monkey and human tissues examined by immunoblot and/or immunohistochemistry, BCATm and BCATc were distributed in patterns very similar to those found in the rat. The results show that BCATm is in a position to regulate BCAA availability as protein precursors and anabolic signals in secretory portions of the digestive and other organ systems. The unique expression of BCATc in neurons of the peripheral nervous system, without coexpression of BCKD, raises new questions about the physiological function of this BCAT isozyme.

The Catabolism of Amino Acids to Long Chain and Complex Alcohols in Saccharomyces cerevisiae

The catabolism of phenylalanine to 2-phenylethanol and of tryptophan to tryptophol were studied by (13)C NMR spectroscopy and gas chromatography-mass spectrometry. Phenylalanine and tryptophan are first deaminated (to 3-phenylpyruvate and 3-indolepyruvate, respectively) and then decarboxylated. This decarboxylation can be effected by any of Pdc1p, Pdc5p, Pdc6p, or Ydr380wp; Ydl080cp has no role in the catabolism of either amino acid. We also report that in leucine catabolism Ydr380wp is the minor decarboxylase. Hence, all amino acid catabolic pathways studied to date use a subtly different spectrum of decarboxylases from the five-membered family that comprises Pdc1p, Pdc5p, Pdc6p, Ydl080cp, and Ydr380wp. Using strains containing all possible combinations of mutations affecting the seven AAD genes (putative aryl alcohol dehydrogenases), five ADH genes, and SFA1, showed that the final step of amino acid catabolism (conversion of an aldehyde to a long chain or complex alcohol) can be accomplished by any one of the ethanol dehydrogenases (Adh1p, Adh2p, Adh3p, Adh4p, Adh5p) or by Sfa1p (formaldehyde dehydrogenase.)

Early steps of metabolism evolution inferred by cladistic analysis of amino acid catabolic pathways

Among abiotic molecules available in primitive environments, free amino acids are good candidates as the first source of energy and molecules for early protocells. Amino acid catabolic pathways are likely to be one of the very first metabolic pathways of life. Among them, which ones were the first to emerge? A cladistic analysis of catabolic pathways of the sixteen aliphatic amino acids and two portions of the Krebs cycle is performed using four criteria of homology. The cladogram shows that the earliest pathways to emerge are not portions of the Krebs cycle but catabolisms of aspartate, asparagine, glutamate, glutamine, proline, arginine. Earliest enzymatic catabolic functions were deaminations and transaminations. Later on appeared enzymatic decarboxylations. The consensus tree allows to propose four time spans for catabolism development and corroborates the views of Cordón in 1990 about the evolution of catabolism.

Production of cheese flavour compounds derived from amino acid catabolism by Propionibacterium freudenreichii

The catabolism of amino acids by cheese micro-organisms results in the production of various volatile flavour compounds. It was recently shown to be a rate-limiting factor in the formation of cheese flavour, leading to an increased interest in elucidating the pathways and the flora involved. This paper reviews the ability of propionibacteria (PAB) to produce flavour compounds deriving from branched-chain, aromatic and sulphur-containing amino acids. In culture media, PAB produced volatile compounds derived from Leu, Ile, Met and Phe. In cheese, the presence of PAB is positively correlated to the amount of acids, alcohols and/or aldehydes derived from Leu or Ile. The metabolic pathways of amino acid conversion to flavour compounds by PAB have been only partly elucidated. Aminotransferase(s) catalyse the first step of conversion of branched-chain, aromatic amino acids and methionine, with a higher activity for branched-chain amino acids. The α-keto acids resulting from transamination are further degraded to various compounds by resting cells of PAB. So α-ketoisocaproic acid, derived from Leu, is essentially converted to isovaleric acid by a ketoacid dehydrogenase complex; phenylpyruvic acid, derived from Phe, is converted to phenyllactic acid, phenylacetic acid, benzoic acid and benzaldehyde. Methionine can also be directly degraded by α, γ -elimination, leading to methanethiol. The amino acid catabolism pathways in PAB share similar- ities with those of lactic acid bacteria but PAB seem to produce higher amounts of branched-chain ac- ids, which are important flavour compounds in cheese. propionibacteria / flavour compound / amino acid / catabolism / cheese ripening

The Escherichia coli starvation gene cstC is involved in amino acid catabolism.

Escherichia coli strains mutant in the starvation gene cstC grow normally in a mineral salts medium but are impaired in utilizing amino acids as nitrogen sources. They are also compromised in starvation survival, where amino acid catabolism is important. The cstC gene encodes a 406-amino-acid protein that closely resembles the E. coli ArgD protein, which is involved in arginine biosynthesis. We postulate that CstC is a counterpart of ArgD in an amino acid catabolic pathway. The cstC upstream region contains several regulatory consensus sequences. Both sigmaS and sigma54 promoters are probably involved in cstC transcription and appear to compete with each other, presumably to match cstC expression to the cellular amino acid catabolic needs.