Lysine biosynthesis in Saccharomyces. Conversion of -aminoadipate into -aminoadipic -semialdehyde.
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ABSTRACT:
SUBMITTER: Sinha AK
PROVIDER: S-EPMC1178178 | biostudies-other | 1971 Dec
REPOSITORIES: biostudies-other
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Project description:The measurements of lysine metabolites provide valuable information for the rapid diagnosis of pyridoxine-dependent epilepsy (PDE). Here, we aimed to develop a sensitive method to simultaneously quantify multiple lysine metabolites in PDE, including α-aminoadipic semialdehyde (a-AASA), piperideine-6-carboxylate (P6C), pipecolic acid (PA) and α-aminoadipic acid (α-AAA) in plasma, serum, dried blood spots (DBS), urine and dried urine spots (DUS). Fifteen patients with molecularly confirmed PDE were detected using liquid chromatography-mass spectrometry (LC-MS/MS) method. Compared to the control groups, the concentrations of a-AASA, P6C and the sum of a-AASA and P6C (AASA-P6C) in all types of samples from PDE patients were markedly elevated. The PA and a-AAA concentrations ranges overlapped partially between PDE patients and control groups. The concentrations of all the analytes in plasma and serum, as well as in urine and DUS were highly correlated. Our study provided more options for the diverse sample collection in the biochemical tests according to practical requirements. With treatment modality of newly triple therapy investigated, biomarker study might play important role not only on diagnosis but also on treatment monitoring and fine tuning the diet. The persistently elevated analytes with good correlation between plasma and DBS, as well as urine and DUS made neonatal screening using DBS and DUS possible.
| S-EPMC6684619 | biostudies-literature
Project description:Pathological levels of oxidative stress (OS) have been implicated in many diseases including diabetes mellitus, neurodegenerative diseases, inflammatory diseases, atherosclerosis, and cancer. Studies of oxidative stress are however complicated by the low concentration of oxidation products. To resolve this problem, we tested a new derivative of aminoadipic semialdehyde (Fmoc-Aea-OH) in the solid-phase synthesis of carbonylated peptides. We prepared a series of peptides with free and acetylated N-terminal amino groups using the Fmoc-Aea-OH reagent. LC-MS, ESI-MS, and MS/MS spectra confirmed the sequences of the modified peptides, although the LC-MS and ESI-MS spectra were dominated by signals corresponding to dehydration products. NMR studies of acetylated products revealed that the dominant product formed in this reaction contains a 1,2,3,4-tetrahydropyridine-2-carboxylic acid residue. Another side reaction in this system was the cleavage of the amide bond between the Aea residue and the amino acid moiety preceding it resulting in the formation of a side product with a six-membered ring at the N-terminus (2,3,4,5-tetrahydropyridine-2-carboxylic acid residue). We found that, depending on the peptide sequence, one of those side products is predominant. Our work suggests new methods for the solid-state synthesis of peptides containing unnatural amino acids.
| S-EPMC6041278 | biostudies-literature
Project description:Purpose: To evaluate the effects of a single oral dose of pyridoxine on lysine metabolites including α-aminoadipic semialdehyde (a-AASA), piperideine-6-carboxylate (P6C), the sum of AASA and P6C (AASA-P6C), pipecolic acid (PA), and α-aminoadipic acid (α-AAA) in PDE patients. Methods: The lysine metabolites of 15 patients with molecularly confirmed PDE were detected before and 4 h after taking a single oral dose of pyridoxine, respectively, using liquid chromatography-mass spectrometry (LC-MS/MS) method. Five types of samples were freshly prepared, including plasma, serum, dried blood spots (DBS), urine, and dried urine spots (DUS). Results: All the patients had been treated with long-term oral pyridoxine for several months to years, with doses of 30-360 mg/d. The concentrations of a-AASA, P6C, AASA-P6C, PA, and a-AAA before and after taking a single oral dose of pyridoxine for the same analyte detected in the same type of sample varied among patients. The mean concentrations increased in almost all the metabolites after taking an oral dose of pyridoxine, with or without statistical significance. Whereas, the metabolites concentrations might increase or decrease among different patients, or in different samples of the same patient, without a regular tendency. There was no statistical correlation between the concentrations before and after taking pyridoxine in the same type of sample for most metabolites. Conclusions: No obvious relationship between the metabolite levels or concentration differences and the age, pyridoxine dose (a single oral dose and long-term maintenance dose), duration of treatment, or neurodevelopmental phenotype was found at present study. The large individual differences among patients, probably affected by various genotypes, leading to quite different effects of pyridoxine on the change degree of metabolites concentrations. Our study suggested that long-term pyridoxine treatment could control seizures rather than getting toxic lysine metabolites such as a-AASA and P6C back to normal. In the future, more therapies should be focused to alleviate the metabolites accumulation and further improve the prognosis of PDE.
| S-EPMC6718124 | biostudies-literature
Project description:The site-specific identification of ?-aminoadipic semialdehyde (AAS) and ?-glutamic semialdehyde (GGS) residues in proteins is reported. Semialdehydic protein modifications result from the metal-catalyzed oxidation of Lys or Arg and Pro residues, respectively. Most of the analytical methods for the analysis of protein carbonylation measure change to the global level of carbonylation and fail to provide details regarding protein identity, site, and chemical nature of the carbonylation. In this work, we used a targeted approach, which combines chemical labeling, enrichment, and tandem mass spectrometric analysis, for the site-specific identification of AAS and GGS sites in proteins. The approach is applied to in vitro oxidized glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and an untreated biological sample, namely cardiac mitochondrial proteins. The analysis of GAPDH resulted in the site-specific identification of two AAA and four GGS residues. Computational evaluation of the identified AAS and GGS sites in GAPDH indicated that these sites are located in flexible regions, show high solvent accessibility values, and are in proximity with possible metal ion binding sites. The targeted proteomic analysis of semialdehydic modifications in cardiac mitochondria yielded nine AAS modification sites which were unambiguously assigned to distinct lysine residues in the following proteins: ATP/ATP translocase isoforms 1 and 2, ubiquinol cytochrome-c reductase core protein 2, and ATP synthase ?-subunit.
| S-EPMC3071856 | biostudies-literature
Project description:The first two steps in the mammalian lysine-degradation pathway are catalyzed by lysine-ketoglutarate reductase and saccharopine dehydrogenase, respectively, resulting in the conversion of lysine to alpha-aminoadipic semialdehyde. Defects in one or both of these activities result in familial hyperlysinemia, an autosomal recessive condition characterized by hyperlysinemia, lysinuria, and variable saccharopinuria. In yeast, lysine-ketoglutarate reductase and saccharopine dehydrogenase are encoded by the LYS1 and LYS9 genes, respectively, and we searched the available sequence databases for their human homologues. We identified a single cDNA that encoded an apparently bifunctional protein, with the N-terminal half similar to that of yeast LYS1 and with the C-terminal half similar to that of yeast LYS9. This bifunctional protein has previously been referred to as "alpha-aminoadipic semialdehyde synthase," and we have tentatively designated this gene "AASS." The AASS cDNA contains an open reading frame of 2,781 bp predicted to encode a 927-amino-acid-long protein. The gene has been sequenced and contains 24 exons scattered over 68 kb and maps to chromosome 7q31.3. Northern blot analysis revealed the presence of several transcripts in all tissues examined, with the highest expression occurring in the liver. We sequenced the genomic DNA from a single patient with hyperlysinemia (JJa). The patient is the product of a consanguineous mating and is homozygous for an out-of-frame 9-bp deletion in exon 15, which results in a premature stop codon at position 534 of the protein. On the basis of these and other results, we propose that AASS catalyzes the first two steps of the major lysine-degradation pathway in human cells and that inactivating mutations in the AASS gene are a cause of hyperlysinemia.
| S-EPMC1378037 | biostudies-other
Project description:An aspartate kinase-deficient mutant of Thermus thermophilus, AK001, was constructed. The mutant strain did not grow in a minimal medium, suggesting that T. thermophilus contains a single aspartate kinase. Growth of the mutant strain was restored by addition of both threonine and methionine, while addition of lysine had no detectable effect on growth. To further elucidate the lysine biosynthetic pathway in T. thermophilus, lysine auxotrophic mutants of T. thermophilus were obtained by chemical mutagenesis. For all lysine auxotrophic mutants, growth in a minimal medium was not restored by addition of diaminopimelic acid, whereas growth of two mutants was restored by addition of alpha-aminoadipic acid, a precursor of lysine in biosynthetic pathways of yeast and fungi. A BamHI fragment of 4.34 kb which complemented the lysine auxotrophy of a mutant was cloned. Determination of the nucleotide sequence suggested the presence of homoaconitate hydratase genes, termed hacA and hacB, which could encode large and small subunits of homoaconitate hydratase, in the cloned fragment. Disruption of the chromosomal copy of hacA yielded mutants showing lysine auxotrophy which was restored by addition of alpha-aminoadipic acid or alpha-ketoadipic acid. All of these results indicated that in T. thermophilus, lysine was not synthesized via the diaminopimelic acid pathway, believed to be common to all bacteria, but via a pathway using alpha-aminoadipic acid as a biosynthetic intermediate.
| S-EPMC93567 | biostudies-literature
Project description:As an indispensable essential amino acid in the human body, lysine is extremely rich in edible mushrooms. The α-aminoadipic acid (AAA) pathway is regarded as the biosynthetic pathway of lysine in higher fungal species in Agaricomycetes. However, there is no deep understanding about the molecular evolutionary relationship between lysine biosynthesis and species in Agaricomycetes. Herein, we analyzed the molecular evolution of lysine biosynthesis in Agaricomycetes. The phylogenetic relationships of 93 species in 34 families and nine orders in Agaricomycetes were constructed with six sequences of LSU, SSU, ITS (5.8 S), RPB1, RPB2, and EF1-α datasets, and then the phylogeny of enzymes involved in the AAA pathway were analyzed, especially homocitrate synthase (HCS), α-aminoadipate reductase (AAR), and saccharopine dehydrogenase (SDH). We found that the evolution of the AAA pathway of lysine biosynthesis is consistent with the evolution of species at the order level in Agaricomycetes. The conservation of primary, secondary, predicted tertiary structures, and substrate-binding sites of the enzymes of HCS, AAR, and SDH further exhibited the evolutionary conservation of lysine biosynthesis in Agaricomycetes. Our results provide a better understanding of the evolutionary conservation of the AAA pathway of lysine biosynthesis in Agaricomycetes.
| S-EPMC8779187 | biostudies-literature
Project description:Herbicides with novel modes of action are urgently needed to safeguard global agricultural industries against the damaging effects of herbicide-resistant weeds. We recently developed the first herbicidal inhibitors of lysine biosynthesis, which provided proof-of-concept for a promising novel herbicide target. In this study, we expanded upon our understanding of the mode of action of herbicidal lysine biosynthesis inhibitors. We previously postulated that these inhibitors may act as proherbicides. Here, we show this is not the case. We report an additional mode of action of these inhibitors, through their inhibition of a second lysine biosynthesis enzyme, and investigate the molecular determinants of inhibition. Furthermore, we extend our herbicidal activity analyses to include a weed species of global significance.
| S-EPMC9208756 | biostudies-literature
Project description:Expression of amino acid biosynthesis genes in bacteria is often repressed when abundant supplies of the cognate amino acid are available. Repression of the Bacillus subtilis lysC gene by lysine was previously shown to occur at the level of premature termination of transcription. In this study we show that lysine directly promotes transcription termination during in vitro transcription with B. subtilis RNA polymerase and causes a structural shift in the lysC leader RNA. We find that B. subtilis lysC is a member of a large family of bacterial lysine biosynthesis genes that contain similar leader RNA elements. By analogy with related regulatory systems, we designate this leader RNA pattern the "L box." Genes in the L box family from Gram-negative bacteria appear to be regulated at the level of translation initiation rather than transcription termination. Mutations of B. subtilis lysC that disrupt conserved leader features result in loss of lysine repression in vivo and loss of lysine-dependent transcription termination in vitro. The identification of the L box pattern also provides an explanation for previously described mutations in both B. subtilis and Escherichia coli lysC that result in lysC overexpression and resistance to the lysine analog aminoethylcysteine. The L box regulatory system represents an example of gene regulation using an RNA element that directly senses the intracellular concentration of a small molecule.
| S-EPMC218712 | biostudies-literature
Project description:A four-step flavanone biosynthetic pathway was constructed and introduced into Saccharomyces cerevisiae. The recombinant yeast strain was fed with phenylpropanoid acids and produced the flavanones naringenin and pinocembrin 62 and 22 times more efficiently compared to previously reported recombinant prokaryotic strains. Microbial biosynthesis of the flavanone eriodictyol was also achieved.
| S-EPMC1214632 | biostudies-literature