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Michaelis-Menten kinetic parameters of PydD (error values reported are the standard errors for the fits).
Source publication
The reductive pyrimidine catabolic pathway is the most widespread pathway for pyrimidine degradation in bacteria, enabling assimilation of nitrogen for growth. This pathway, which has been studied in several bacteria including Escherichia coli B, releases only one utilizable nitrogen atom from each molecule of uracil, while the other nitrogen atom...
Contexts in source publication
Context 1
... The resulting consensus tree ( Fig. 5) was rendered using the web-based program iTOL 21 . Table 1. With β-alanine as the amine donor, the highest activity was obtained with 2-oxoglutarate as the amine acceptor ( Figure S5, S6), suggesting that 2-oxoglutarate is the physiological substrate for PydD. ...Context 2
... β-alanine as the amine donor, the highest activity was obtained with 2-oxoglutarate as the amine acceptor ( Figure S5, S6), suggesting that 2-oxoglutarate is the physiological substrate for PydD. The apparent k cat /K M for pyruvate was 63-fold lower than that for 2-oxoglutarate (Table 1). Under the same reaction conditions, the activity was the lowest for oxaloacetate ( Figure S6), and its apparent k cat /K M was not determined. ...Similar publications
Cancers disrupt host homeostasis in various manners but the identity of host factors underlying such disruption remains largely unknown. Here we show that nicotinamide-N-methyltransferase (NNMT) is a host factor that mediates metabolic dysfunction in the livers of cancer-bearing mice. Multiple solid cancers distantly increase expression of Nnmt and...
Citations
... Moreover, B. smithii S-2710M can utilize LG at the same level as glucose without being inhibited by catabolic products, a finding that contributes to the efficient application of LG (Kuritani et al., 2020). Accumulation of 3-hydroxypropionic acid was detected in B. smithii CGMCC 1.3752 grown in a specific medium with uracil or uridine as the sole nitrogen source, findings that also extend the diversity of reductive pyrimidine catabolic pathways and pyrimidine-degrading enzymes (Yin et al., 2019). DNA polymerases play a vital role in biological research, and the development of a new generation of DNA polymerases could save money and time and lead to more applications. ...
Bacillus smithii is a thermophilic Bacillus that can be isolated from white wine, hot spring soil, high‐temperature compost, and coffee grounds, with various biofunctions and wide applications. It is resistant to both gastric acid and high temperature, which makes it easier to perform probiotic effects than traditional commercial probiotics, so it can maintain good vitality during food processing and has great application prospects. This paper starts with the taxonomy and genetics and focuses on aspects, including genetic transformation, functional enzyme production, waste utilization, and application in the field of food science as a potential probiotic. According to available studies during the past 30 years, we considered that B. smithii is a novel class of microorganisms with a wide range of functional enzymes such as hydrolytic enzymes and hydrolases, as well as resistance to pathogenic bacteria. It is available in waste degradation, organic fertilizer production, the feed and chemical industries, the pharmaceutical sector, and food fortification. Moreover, B. smithii has great potentials for applications in the food industry, as it presents high resistance to the technological processes that guarantee its health benefits. It is also necessary to systematically evaluate the safety, flavor, and texture of B. smithii and explore its biological mechanism of action, which is of great value for further application in multiple fields, especially in food and medicine.
... Alternatively, this may indicate that oxaloacetate undergoes preceding oxidative decarboxylation to 3-oxopropanoic acid, which in turn reacts with pyridoxamine to form β-alanine. Such a reaction would mimic a reversed β-alanine degradation pathway [79]. ...
A protometabolic approach to the origins of life assumes that the conserved biochemistry of metabolism has direct continuity with prebiotic chemistry. One of the most important amino acids in modern biology is aspartic acid, serving as a nodal metabolite for the synthesis of many other essential biomolecules. Aspartate’s prebiotic synthesis is complicated by the instability of its precursor, oxaloacetate. In this paper, we show that the use of the biologically relevant cofactor pyridoxamine, supported by metal ion catalysis, is sufficiently fast to offset oxaloacetate’s degradation. Cu2+-catalysed transamination of oxaloacetate by pyridoxamine achieves around a 5% yield within 1 h, and can operate across a broad range of pH, temperature, and pressure. In addition, the synthesis of the downstream product β-alanine may also take place in the same reaction system at very low yields, directly mimicking an archaeal synthesis route. Amino group transfer supported by pyridoxal is shown to take place from aspartate to alanine, but the reverse reaction (alanine to aspartate) shows a poor yield. Overall, our results show that the nodal metabolite aspartate and related amino acids can indeed be synthesised via protometabolic pathways that foreshadow modern metabolism in the presence of the simple cofactor pyridoxamine and metal ions.
... β-Alanine can be biologically synthesized Shilong Hu and Mingyue Fei contributed equally to this work. by several pathways, including degradation of pyrimidine or polyamine, multistep catalysis with propionate, and decarboxylation of L-aspartic acid (L-ASP) (Jr 1980;Li et al. 2017;Parthasarathy et al. 2019;Wang et al. 2021a;White et al. 2001;Yin et al. 2019). Traditionally, β-ALA was synthesized through chemical reactions (Dean, 1943;Ford 1945), and whole-cell catalysis (Li et al. 2018;Piao et al. 2019;Toshihiro O. et al., 1998). ...
β-alanine has been used in food and pharmaceutical industries. Although Escherichia coli Nissle 1917 (EcN) is generally considered safe and engineered as living therapeutics, engineering EcN for producing industrial metabolites has rarely been explored. Here, by protein and metabolic engineering, EcN was engineered for producing β-alanine from glucose. First, an aspartate-α-decarboxylase variant ADCK43Y with improved activity was identified and over-expressed in EcN, promoting the titer of β-alanine from an undetectable level to 0.46 g/L. Second, directing the metabolic flux towards L-aspartate increased the titer of β-alanine to 0.92 g/L. Third, the yield of β-alanine was elevated to 1.19 g/L by blocking conversion of phosphoenolpyruvate to pyruvate, and further increased to 2.14 g/L through optimizing culture medium. Finally, the engineered EcN produced 11.9 g/L β-alanine in fed-batch fermentation. Our work not only shows the potential of EcN as a valuable industrial platform, but also facilitates production of β-alanine via fermentation.
Key points
• Escherichia coli Nissle 1917 (EcN) was engineered as a β-alanine producing cell factory
• Identification of a decarboxylase variant ADCK43Y with improved activity
• Directing the metabolic flux to L-ASP and expressing ADCK43Y elevated the titer of β-alanine to 11.9 g/L
... The contents of xanthine, hypoxanthine, guanosine, guanine, adenosine, adenine, 2'-deoxyadenosine, (R) (-)-allantoin, uracil, thymine and cytidine were affected, which indicated that the purine and pyrimidine metabolism pathways were affected. This was also reported previously by Liang et al. [27], and Yin et al. [28] observed variations in purine and pyrimidine metabolism when bacteria were fed diets with different protein contents. The enhanced purine and pyrimidine metabolism of the high-protein group was attributed to the increased MCP synthesis in the rumen [29]. ...
This experiment was aimed to compare the effects of two diets with different protein content on the growth performance, immune indexes, rumen fermentation characteristics and plasma metabolomics of growing yak in the cold season. A total of 24, 2-year-old healthy yaks with similar body weight (142.9 ± 3.56 kg) were randomly allocated to two isoenergetic diets with different protein content (10 vs 14%) according to a non-paired experimental design, and the protein of the diets was increased by increasing soybean meal, rapeseed meal and cottonseed meal. The growth performance experiment lasted 56 days. Four days before the end of the growth experiment, the digestion trial was conducted, and the rumen fluid and plasma was collected for measurement. The results showed that the average daily feed intake (p
... In contrast, metabolic functions in the control condition were highly associated with nitrogen, beta-alanine, fatty acid, and sulfur metabolisms and benzoate degradation. These metabolic pathways are associated with several organic degradation and nutrient metabolism outcomes, which suggests that the pathways involved in organic degradation and nitrogen and sulfur availabilities are dominant around the plant roots during control conditions (Singh and Schwan, 2011;Yin et al., 2019). Nitrogen and sulfur metabolisms are important for the formation of amino acids, particularly cysteine, which is then utilized in protein synthesis and mediate nutrient availability (e.g., nitrogen and sulfur) for plant growth (Ren et al., 2017;Singh and Schwan, 2011). ...
Bioponics has the potential to recover nutrients from organic waste streams, such as chicken manure and digestate with high volatile fatty acid (VFA) contents through crop production. Acetic acid, a dominant VFA, was supplemented weekly (0, 500, 1000, and 1500 mg/L) in a chicken manure-based bioponic system, and its effect on the performance of bioponics (e.g., plant yield and nitrogen and phosphorus availabilities) was examined. Microbial communities were analyzed using 16S rRNA gene sequencing, and the functional gene abundances were predicted using PICRUSt. Although acetic acid negatively affected plant yield, no significant difference (p > 0.05) was noted in the average nitrogen or phosphorus concentration. In terms of nutrient recovery, the bioponic systems still functioned well, although higher concentrations of acetic acid decreased plant yield and altered the bacterial communities in plant roots and chicken manure sediments. These data suggest that an acetic acid concentration of < 500 mg/L or a longer loading interval is recommended for the effective operation of chicken manure and digestate-based bioponics.
... There is evidence of an adaptive response of Escherichia-Shigella members to long-term nitrogen starvation in anaerobic environments, which requires allantoin degradation (Switzer et al. 2020). It is worth noting that E. coli can obtain four nitrogen atoms from purines, while it can only gain one nitrogen atom from each uracil molecule through the reductive pyrimidine catabolic pathway (Salway 2018;Yin et al. 2019). We hypothesized that the increased activity of xanthine dehydrogenase and the enzymes of ureide synthesis in tophaceous gout patients is likely an adaptive process of the gut microbiome, which may be attributable to more than 8 years of allopurinol treatment. ...
Objective
To evaluate the taxonomic composition of the gut microbiome in gout patients with and without tophi formation, and predict bacterial functions that might have an impact on urate metabolism.
Methods
Hypervariable V3–V4 regions of the bacterial 16S rRNA gene from fecal samples of gout patients with and without tophi (n = 33 and n = 25, respectively) were sequenced and compared to fecal samples from 53 healthy controls. We explored predictive functional profiles using bioinformatics in order to identify differences in taxonomy and metabolic pathways.
Results
We identified a microbiome characterized by the lowest richness and a higher abundance of Phascolarctobacterium , Bacteroides , Akkermansia , and Ruminococcus_gnavus_group genera in patients with gout without tophi when compared to controls. The Proteobacteria phylum and the Escherichia-Shigella genus were more abundant in patients with tophaceous gout than in controls. Fold change analysis detected nine genera enriched in healthy controls compared to gout groups ( Bifidobacterium, Butyricicoccus, Oscillobacter, Ruminococcaceae_UCG_010, Lachnospiraceae_ND2007_group, Haemophilus, Ruminococcus_1, Clostridium_sensu_stricto_1, and Ruminococcaceae_UGC_013 ). We found that the core microbiota of both gout groups shared Bacteroides caccae , Bacteroides stercoris ATCC 43183 , and Bacteroides coprocola DSM 17136 . These bacteria might perform functions linked to one-carbon metabolism, nucleotide binding, amino acid biosynthesis, and purine biosynthesis. Finally, we observed differences in key bacterial enzymes involved in urate synthesis, degradation, and elimination.
Conclusion
Our findings revealed that taxonomic variations in the gut microbiome of gout patients with and without tophi might have a functional impact on urate metabolism.
... The proposed roles of HpfD homologs in metabolism of the C3 sulfonate 3-HPS and structurally related C4 carboxylates GABA and 4-HB are analogous to the roles previously demonstrated for the M-ADH enzyme TauF in metabolism of the C2 sulfonate taurine and 2-hydroxyethylsulfonate (isethionate) and C3 carboxylates β-alanine and 3-hydroxypropionate. [18,19] In conclusion, our biochemical investigation of HpfD revealed an unexpected degree of substrate promiscuity, underscoring the inter-relationship between enzymes involved in sulfonate and carboxylate metabolism. Microbiome-derived carboxylates like GABA and succinate have known physiological effects on the human host, though the effects of their sulfonate isosteres are unknown. ...
Sulfoquinovose is the polar headgroup of plant sulfolipids and is a globally abundant organosulfur compound, and its degradation by bacteria is an important component of the sulfur cycle. Sulfoquinovose degradation by certain bacteria, including Escherichia coli, produces dihydroxypropanesulfonate (DHPS), which is further converted by anaerobic bacteria into 3‐hydroxypropanesulfonate (3‐HPS), through the catalytic action of DHPS dehydratase (a member of the glycyl radical enzyme family), and sulfopropionaldehyde reductase HpfD (a member of the metal‐dependent alcohol dehydrogenase family). Here we report biochemical investigation of Hungatella hathewayi HpfD. In addition to 3‐HPS, HpfD also displayed high catalytic activities for NAD⁺‐dependent oxidation of 4‐hydroxybutanesulfonate (4‐HBS) and γ‐hydroxybutyrate (GHB). The highest activity was obtained with Fe²⁺ or Mn²⁺ as the divalent metal cofactor. Bioinformatics studies suggest that, in addition to DHPS degradation, 3‐HPS and γ‐aminobutyrate (GABA) degradations also involve HpfD homologs.
... In this pathway, uracil is first reduced to dihydrouracil, catalyzed by dihydropyrimidine reductase (PydA) [2,3]. This de-aromatization of the pyrimidine ring facilitates subsequent hydrolysis by dihydropyrimidinase (PydB) and ureidopropionase (PydC), releasing ammonia and β-alanine for use as nitrogen and / or carbon sources for growth [2][3][4][5]. ...
... To investigate the occurrence of PydAc in bacteria, a SSN of 3419 candidate PydA sequences was constructed (Fig. 5), and the gene clusters were examined for the presence of a pyridine nucleotide oxidoreductase domain (Pyr_redox_2, PF07992), indicative in theory of the ability to use NAD(P)H as a reductant. The majority of the PydA sequences (2781 total) contained a Pyr_redox_2 domain protein in an adjacent ORF (Fig. 5a), including several Downloaded from https://portlandpress.com/bioscirep/article-pdf/doi/10.1042/BSR20201642/884753/bsr-2020-1642-t.pdf by guest on 18 June 2020 organisms, in which the Pyd pathway was previously studied (Brevibacillus agri [3], Bacillus smithii [4], Bacillus megaterium [5], Pseudomonas putida [19]). A further 29 sequences contained a Pyr_redox_2 domain within the same polypeptide. ...
Dihydropyrimidine dehydrogenase (PydA) catalyzes the first step of the reductive pyrimidine degradation (Pyd) pathway in bacteria and eukaryotes, enabling pyrimidines to be utilized as substrates for growth. PydA homologs studied to date catalyze the reduction of uracil to dihydrouracil, coupled to the oxidation of NAD(P)H. Uracil reduction occurs at a flavin mononucleotide (FMN) site, and NAD(P)H oxidation occurs at a flavin adenine dinucleotide (FAD) site, with two ferredoxin domains thought to mediate inter-site electron transfer. Here we report the biochemical characterization of a Clostridial PydA homolog (PydAc) from a Pyd gene cluster in the strict anaerobic bacterium Clostridium chromiireducens. PydAc lacks the FAD domain, and instead is able to catalyze uracil reduction using reduced methyl viologen or reduced ferredoxin as the electron source. Homologs of PydAc are present in Pyd gene clusters in many strict anaerobic bacteria, which use reduced ferredoxin as an intermediate in their energy metabolism.
... However, with a changing gene neighborhood and regulation, IsfD and related SDR 3-hydroxyacid dehydrogenases may be involved in other pathways as well, for example, for the detoxification of malonic semialdehyde. In this connection, the metabolic sequence for the deamination of β-alanine via malonic semialdehyde and 3-hydroxypropionate catalyzed by pyridoxal phosphate-dependent transaminase PydD and reductase PydE, respectively, has recently been elucidated in the bacterium Lysinibacillus massiliensis [26]. Like SarD and TauF, PydE is a metal-depending dehydrogenase not related to the SDR superfamily. ...
In last year's issue 4 of Biochemical Journal, Zhou et al. (Biochem J. 476, 733–746) kinetically and structurally characterized the reductase IsfD from Klebsiella oxytoca that catalyzes the reversible reduction in sulfoacetaldehyde to the corresponding alcohol isethionate. This is a key step in detoxification of the carbonyl intermediate formed in bacterial nitrogen assimilation from the α-aminoalkanesulfonic acid taurine. In 2019, the work on sulfoacetaldehyde reductase IsfD was the exciting start to a quite remarkable series of articles dealing with structural elucidation of proteins involved in taurine metabolism as well as the discovery of novel degradation pathways in bacteria.
... Recently, we reported an extended Pyd pathway in Lysinibacillus massiliensis, involving a -alanine:2-oxoglutarate aminotransferase (PydD1) and an NAD(P)Hdependent malonic semialdehyde reductase (PydE) (8). PydD1 generates glutamate, an intermediate in nitrogen metabolism, thus enabling the assimilation of the second pyrimidine nitrogen atom. ...
... PydD1 generates glutamate, an intermediate in nitrogen metabolism, thus enabling the assimilation of the second pyrimidine nitrogen atom. PydE converts the toxic intermediate malonic semialdehyde into 3-hydroxypropionate (3-HP), which is excreted into the medium (8). In addition to being incorporated into bacterial nitrogen metabolism, -alanine could also in principle be incorporated into bacterial carbon metabolism, allowing pyrimidines to be used as a carbon source for growth. ...
... In addition to PydABC, many Bacillales bacteria contain a -alanine:pyruvate aminotransferase (PydD2) and malonic semialdehyde dehydrogenase (MSDH), while many Clostridiales bacteria contain a -alanine:2oxoglutarate aminotransferase (PydD1) and malonic semialdehyde reductase (PydE). (B) Comparison of the two extended reductive pyrimidine degradation pathways, involving either PydD1 and PydE as previously reported (8) or PydD2 and MSDH as described in this study. ...
Bacteria utilize diverse biochemical pathways for the degradation of the pyrimidine ring. The function of the pathways studied to date has been the release of nitrogen for assimilation. The most widespread of these pathways is the reductive pyrimidine catabolic pathway, which converts uracil into ammonia, carbon dioxide and β-alanine. Here we report the characterization of a β-alanine:pyruvate aminotransferase (PydD2), and a NAD ⁺ -dependent malonic semialdehyde dehydrogenase (MSDH), from a reductive pyrimidine catabolism gene cluster in Bacillus megaterium . Together, these enzymes convert β-alanine into acetyl-CoA, a key intermediate in carbon and energy metabolism. We demonstrate the growth of B. megaterium in defined medium with uracil as its sole carbon and energy source. Homologs of PydD2 and MSDH are found in association with reductive pyrimidine pathway genes in many Gram positive bacteria in the order Bacillales. Our study provides a basis for further investigations of the utilization of pyrimidines as a carbon and energy source by bacteria.
Importance Pyrimidine has wide occurrence in natural environments, where bacteria use it as nitrogen and carbon source for growth. Detailed biochemical pathways have been investigated with focus mainly on nitrogen assimilation in the past decades. Here we report the discovery and characterization of two important enzymes, PydD2 and MSDH, which constitute an extension for the reductive pyrimidine catabolic pathway. These two enzymes, prevalent in Bacillales based on our bioinformatics studies, allow stepwise conversion of β-alanine, a previous “end product” of the reductive pyrimidine degradation pathway to acetyl-CoA, as carbon and energy source.
