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Microbiome May 2024Left ventricular diastolic dysfunction (LVDD) is an important precursor of heart failure (HF), but little is known about its relationship with gut dysbiosis and...
BACKGROUND
Left ventricular diastolic dysfunction (LVDD) is an important precursor of heart failure (HF), but little is known about its relationship with gut dysbiosis and microbial-related metabolites. By leveraging the multi-omics data from the Hispanic Community Health Study/Study of Latinos (HCHS/SOL), a study with population at high burden of LVDD, we aimed to characterize gut microbiota associated with LVDD and identify metabolite signatures of gut dysbiosis and incident LVDD.
RESULTS
We included up to 1996 Hispanic/Latino adults (mean age: 59.4 years; 67.1% female) with comprehensive echocardiography assessments, gut microbiome, and blood metabolome data. LVDD was defined through a composite criterion involving tissue Doppler assessment and left atrial volume index measurements. Among 1996 participants, 916 (45.9%) had prevalent LVDD, and 212 out of 594 participants without LVDD at baseline developed incident LVDD over a median 4.3 years of follow-up. Using multivariable-adjusted analysis of compositions of microbiomes (ANCOM-II) method, we identified 7 out of 512 dominant gut bacterial species (prevalence > 20%) associated with prevalent LVDD (FDR-q < 0.1), with inverse associations being found for Intestinimonas_massiliensis, Clostridium_phoceensis, and Bacteroide_coprocola and positive associations for Gardnerella_vaginali, Acidaminococcus_fermentans, Pseudomonas_aeruginosa, and Necropsobacter_massiliensis. Using multivariable adjusted linear regression, 220 out of 669 circulating metabolites with detection rate > 75% were associated with the identified LVDD-related bacterial species (FDR-q < 0.1), with the majority being linked to Intestinimonas_massiliensis, Clostridium_phoceensis, and Acidaminococcus_fermentans. Furthermore, 46 of these bacteria-associated metabolites, mostly glycerophospholipids, secondary bile acids, and amino acids, were associated with prevalent LVDD (FDR-q < 0.1), 21 of which were associated with incident LVDD (relative risk ranging from 0.81 [p = 0.001, for guanidinoacetate] to 1.25 [p = 9 × 10, for 1-stearoyl-2-arachidonoyl-GPE (18:0/20:4)]). The inclusion of these 21 bacterial-related metabolites significantly improved the prediction of incident LVDD compared with a traditional risk factor model (the area under the receiver operating characteristic curve [AUC] = 0.73 vs 0.70, p = 0.001). Metabolite-based proxy association analyses revealed the inverse associations of Intestinimonas_massilliensis and Clostridium_phoceensis and the positive association of Acidaminococcus_fermentans with incident LVDD.
CONCLUSION
In this study of US Hispanics/Latinos, we identified multiple gut bacteria and related metabolites linked to LVDD, suggesting their potential roles in this preclinical HF entity. Video Abstract.
Topics: Humans; Gastrointestinal Microbiome; Female; Middle Aged; Male; Hispanic or Latino; Ventricular Dysfunction, Left; United States; Dysbiosis; Aged; Bacteria; Metabolome; Echocardiography
PubMed: 38725043
DOI: 10.1186/s40168-024-01797-x -
Molecular Microbiology Jan 1999Glutaconyl-CoA decarboxylase from Acidaminococcus fermentans (clostridal cluster IX), a strict anaerobic inhabitant of animal intestines, uses the free energy of...
Glutaconyl-CoA decarboxylase from Acidaminococcus fermentans (clostridal cluster IX), a strict anaerobic inhabitant of animal intestines, uses the free energy of decarboxylation (delta G(o) approximately -30 kJ mol-1) in order to translocate Na+ from the inside through the cytoplasmic membrane. The proton, which is required for decarboxylation, most probably comes from the outside. The enzyme consists of four different subunits. The largest subunit, alpha or GcdA (65 kDa), catalyses the transfer of CO2 from glutaconyl-CoA to biotin covalently attached to the gamma-subunit, GcdC. The beta-subunit, GcdB, is responsible for the decarboxylation of carboxybiotin, which drives the Na+ translocation (approximate K(m) for Na+ 1 mM), whereas the function of the smallest subunit, delta or GcdD, is unclear. The gene gcdA is part of the 'hydroxyglutarate operon', which does not contain genes coding for the other three subunits. This paper describes that the genes, gcdDCB, are transcribed in this order from a distinct operon. The delta-subunit (GcdD, 12 kDa), with one potential transmembrane helix, probably serves as an anchor for GcdA. The biotin carrier (GcdC, 14 kDa) contains a flexible stretch of 50 amino acid residues (A26-A75), which consists of 34 alanines, 14 prolines, one valine and one lysine. The beta-subunit (GcdB, 39 kDa) comprising 11 putative transmembrane helices shares high amino acid sequence identities with corresponding deduced gene products from Veillonella parvula (80%, clostridial cluster IX), Archaeoglobus fulgidus (61%, Euryarchaeota), Propionigenium modestum (60%, clostridial cluster XIX), Salmonella typhimurium (51%, enterobacteria) and Klebsiella pneumoniae (50%, enterobacteria). Directly upstream of the promoter region of the gcdDCB operon, the 3' end of gctM was detected. It encodes a protein fragment with 73% sequence identity to the C-terminus of the alpha-subunit of methylmalonyl-CoA decarboxylase from V. parvula (MmdA). Hence, it appears that A. fermentans should be able to synthesize this enzyme by expression of gctM together with gdcDCB, but methylmalonyl-CoA decarboxylase activity could not be detected in cell-free extracts. Earlier observations of a second, lower affinity binding site for Na+ of glutaconyl-CoA decarboxylase (apparent K(m) 30 mM) were confirmed by identification of the cysteine residue 243 of GcdB between the putative hellces VII and VIII, which could be specifically protected from alkylation by Na+. The alpha-subunit was purified from an overproducing Escherichia coli strain and was characterized as a putative homotrimer able to catalyse the carboxylation of free biotin.
Topics: Amino Acid Sequence; Base Sequence; Biological Transport; Biotin; Carboxy-Lyases; Cloning, Molecular; DNA, Bacterial; Genes, Bacterial; Gram-Negative Anaerobic Bacteria; Ions; Molecular Sequence Data; Open Reading Frames; Operon; Sodium; Transcription, Genetic
PubMed: 10027965
DOI: 10.1046/j.1365-2958.1999.01189.x -
The Journal of Biological Chemistry Jul 1999The exchange of oxygen atoms between acetate, glutaryl-CoA, and the catalytic glutamate residue in glutaconate CoA-transferase from Acidaminococcus fermentans was...
Oxygen exchange between acetate and the catalytic glutamate residue in glutaconate CoA-transferase from Acidaminococcus fermentans. Implications for the mechanism of CoA-ester hydrolysis.
The exchange of oxygen atoms between acetate, glutaryl-CoA, and the catalytic glutamate residue in glutaconate CoA-transferase from Acidaminococcus fermentans was analyzed using [(18)O(2)]acetate together with matrix-assisted laser desorption/ionization time of flight mass spectrometry of an appropriate undecapeptide. The exchange reaction was shown to be site-specific, reversible, and required both glutaryl-CoA and [(18)O(2)]acetate. The observed exchange is in agreement with the formation of a mixed anhydride intermediate between the enzyme and acetate. In contrast, with a mutant enzyme, which was converted to a thiol ester hydrolyase by replacement of the catalytic glutamate residue by aspartate, no (18)O uptake from H(2)(18)O into the carboxylate was detectable. This result is in accord with a mechanism in which the carboxylate of aspartate acts as a general base in activating a water molecule for hydrolysis of the thiol ester intermediate. This mechanism is further supported by the finding of a significant hydrolyase activity of the wild-type enzyme using acetyl-CoA as substrate, whereas glutaryl-CoA is not hydrolyzed. The small acetate molecule in the substrate binding pocket may activate a water molecule for hydrolysis of the nearby enzyme-CoA thiol ester.
Topics: Bacteria, Anaerobic; Bacterial Proteins; Coenzyme A-Transferases; Enzyme Activation; Glutamic Acid; Hydrolysis; Mass Spectrometry; Oxygen
PubMed: 10409616
DOI: 10.1074/jbc.274.30.20772 -
Microbiome 2015Chronic malnutrition, termed stunting, is defined as suboptimal linear growth, affects one third of children in developing countries, and leads to increased mortality...
BACKGROUND
Chronic malnutrition, termed stunting, is defined as suboptimal linear growth, affects one third of children in developing countries, and leads to increased mortality and poor developmental outcomes. The causes of childhood stunting are unknown, and strategies to improve growth and related outcomes in children have only had modest impacts. Recent studies have shown that the ecosystem of microbes in the human gut, termed the microbiota, can induce changes in weight. However, the specific changes in the gut microbiota that contribute to growth remain unknown, and no studies have investigated the gut microbiota as a determinant of chronic malnutrition.
RESULTS
We performed secondary analyses of data from two well-characterized twin cohorts of children from Malawi and Bangladesh to identify bacterial genera associated with linear growth. In a case-control analysis, we used the graphical lasso to estimate covariance network models of gut microbial interactions from relative genus abundances and used network analysis methods to select genera associated with stunting severity. In longitudinal analyses, we determined associations between these selected microbes and linear growth using between-within twin regression models to adjust for confounding and introduce temporality. Reduced microbiota diversity and increased covariance network density were associated with stunting severity, while increased relative abundance of Acidaminococcus sp. was associated with future linear growth deficits.
CONCLUSIONS
We show that length growth in children is associated with community-wide changes in the gut microbiota and with the abundance of the bacterial genus, Acidaminococcus. Larger cohorts are needed to confirm these findings and to clarify the mechanisms involved.
PubMed: 26106478
DOI: 10.1186/s40168-015-0089-2 -
Journal of Biological Inorganic... May 2008The key enzyme of the fermentation of glutamate by Acidaminococcus fermentans, 2-hydroxyglutaryl-coenzyme A dehydratase, catalyzes the reversible syn-elimination of...
The key enzyme of the fermentation of glutamate by Acidaminococcus fermentans, 2-hydroxyglutaryl-coenzyme A dehydratase, catalyzes the reversible syn-elimination of water from (R)-2-hydroxyglutaryl-coenzyme A, resulting in (E)-glutaconylcoenzyme A. The dehydratase system consists of two oxygen-sensitive protein components, the activator (HgdC) and the actual dehydratase (HgdAB). Previous biochemical and spectroscopic studies revealed that the reduced [4Fe-4S]+ cluster containing activator transfers one electron to the dehydratase driven by ATP hydrolysis, which activates the enzyme. With a tenfold excess of titanium(III) citrate at pH 8.0 the activator can be further reduced, yielding about 50% of a superreduced [4Fe-4S]0 cluster in the all-ferrous state. This is inferred from the appearance of a new Mössbauer spectrum with parameters delta = 0.65 mm/s and deltaE(Q) = 1.51-2.19 mm/s at 140 K, which are typical of Fe(II)S4 sites. Parallel-mode electron paramagnetic resonance (EPR) spectroscopy performed at temperatures between 3 and 20 K showed two sharp signals at g = 16 and 12, indicating an integer-spin system. The X-band EPR spectra and magnetic Mössbauer spectra could be consistently simulated by adopting a total spin S(t) = 4 for the all-ferrous cluster with weak zero-field splitting parameters D = -0.66 cm(-1) and E/D = 0.17. The superreduced cluster has apparent spectroscopic similarities with the corresponding [4Fe-4S]0 cluster described for the nitrogenase Fe-protein, but in detail their properties differ. While the all-ferrous Fe-protein is capable of transferring electrons to the MoFe-protein for dinitrogen reduction, a similar physiological role is elusive for the superreduced activator. This finding supports our model that only one-electron transfer steps are involved in dehydratase catalysis. Nevertheless we discuss a common basic mechanism of the two diverse systems, which are so far the only described examples of the all-ferrous [4Fe-4S]0 cluster found in biology.
Topics: Acidaminococcus; Hydro-Lyases; Iron-Sulfur Proteins; Oxidation-Reduction; Spectrum Analysis
PubMed: 18274792
DOI: 10.1007/s00775-008-0345-z -
European Journal of Biochemistry Dec 19871. The (R)-2-hydroxyglutaryl-CoA dehydratase system from Acidaminococcus fermentans was separated by chromatography of cell-free extracts on Q-Sepharose into two...
1. The (R)-2-hydroxyglutaryl-CoA dehydratase system from Acidaminococcus fermentans was separated by chromatography of cell-free extracts on Q-Sepharose into two components, an activator and the actual dehydratase. The latter enzyme was further purified to homogeneity by chromatography on blue-Sepharose. It is an iron-sulfur protein (Mr 210,000) consisting of two different polypeptides (alpha, Mr 55,000, and beta, Mr 42,000) in an alpha 2 beta 2 structure with probably two [4Fe-4S] centers. After activation this purified enzyme catalysed the dehydration of (R)-2-hydroxyglutarate only in the presence of acetyl-CoA and glutaconate CoA-transferase, demonstrating that the thiol ester and not the free acid is the substrate of the dehydration. The result led to a modification of the hydroxyglutarate pathway of glutamate fermentation. 2. The activation of the dehydratase by the flow-through from Q-Sepharose concentrated by ultrafiltration required NADH, MgCl2, ATP and strict anaerobic conditions. This fraction was designated as Ao. Later when the concentration was performed by chromatography on phenyl-Sepharose, an NADH-independent form of the activator, designated as A*, was obtained. This enzyme, which required only ATP for activation of the dehydratase, was purified further by affinity chromatography on ATP-agarose. It contains neither iron nor inorganic sulfur. A*, as well as the activated dehydratase, were irreversibly inactivated by exposure to air within less than 15 min. The activated dehydratase but not A* was also inactivated by 1 mM hydroxylamine or by 0.1 mM 2,4-dinitrophenol. 3. The (R)-2-hydroxyglutaryl-CoA dehydratase system is closely related the that of (R)-lactoyl-CoA dehydratase from Clostridium propionicum as described by R. D. Kuchta and R. H. Abeles [(1985) J. Biol. Chem. 260, 13,181-13,189].
Topics: Amino Acid Sequence; Bacteria, Anaerobic; Hydro-Lyases; Iron; Iron-Sulfur Proteins; Kinetics; Macromolecular Substances; Metalloproteins; Molecular Sequence Data; Molecular Weight; Molybdenum; Sulfur
PubMed: 3691501
DOI: 10.1111/j.1432-1033.1987.tb13631.x -
FEBS Letters Mar 1997The heterooctameric (alphabeta)4 glutaconate CoA-transferase (EC 2.8.3.12) from the anaerobic bacterium Acidaminococcus fermentans catalyses the transfer of CoASH from... (Comparative Study)
Comparative Study
The heterooctameric (alphabeta)4 glutaconate CoA-transferase (EC 2.8.3.12) from the anaerobic bacterium Acidaminococcus fermentans catalyses the transfer of CoASH from acetyl-CoA to the 1-carboxylate of glutaconate. During this reaction the glutamate residue 54 of the beta-subunit (betaE54) forms a CoA-ester. The single amino acid replacement betaE54D resulted in a drastic change of enzymatic function. The CoA-transferase activity decreased from 140 to less than 0.01 s(-1), whereas the acyl-CoA hydrolase activity increased from less than 0.01 to 16 s(-1). The new enzyme was able to catalyse the hydrolysis of glutaryl-CoA, acetyl-CoA and 3-butenoyl-CoA. Since the mutants betaE54A and betaE54N showed neither acyl-CoA hydrolase nor CoA-transferase activity, it was concluded that the aspartate carboxylate of the mutant betaE54D acted as a general base which facilitated the attack of water at the thiolester carbonyl. Surprisingly, Km for glutaryl-CoA hydrolysis by the mutant (0.7 microM) as compared to CoA-transfer by the wild-type (28 microM) was 40 times lower. A 65 kDa protein, obtained by fusing the genes, gctA-gctB, coding for glutaconate CoA-transferase, retained 30% of the wild-type activity. Comparison of the amino acid sequences of 13 related enzymes demonstrated that Nature already has applied gene fusion in the case of pig heart CoA-transferase and has been using the E --> D mutation for catalysis by a yeast acetyl-CoA hydrolase.
Topics: Amino Acid Sequence; Binding Sites; Coenzyme A-Transferases; Escherichia coli; Genes, Bacterial; Glutamic Acid; Gram-Negative Anaerobic Cocci; Molecular Sequence Data; Mutagenesis, Site-Directed; Mutation; Palmitoyl-CoA Hydrolase; Recombinant Proteins; Substrate Specificity
PubMed: 9089292
DOI: 10.1016/s0014-5793(97)00187-7 -
European Journal of Biochemistry Feb 19931. The primary sodium-ion pump glutaconyl-CoA decarboxylase (GCD) from Acidaminococcus fermentans is composed of four subunits: GCDA, the carboxytransferase (65 kDa),... (Comparative Study)
Comparative Study
Cloning, sequencing and expression of the gene encoding the carboxytransferase subunit of the biotin-dependent Na+ pump glutaconyl-CoA decarboxylase from Acidaminococcus fermentans in Escherichia coli.
1. The primary sodium-ion pump glutaconyl-CoA decarboxylase (GCD) from Acidaminococcus fermentans is composed of four subunits: GCDA, the carboxytransferase (65 kDa), GCDB, the carboxylyase (36 kDa), GCDC, the biotin carrier (24 kDa) and GCDD (14 kDa) of unknown function. A genomic library of A. fermentans was screened with an antiserum raised against whole GCD. A clone giving the strongest reaction in an immunoassay contained a 12-kbp genomic fragment from A. fermentans and was analysed further. An oligonucleotide deduced from the N-terminus of GCDA was used for probing the corresponding gene gcdA. It is 1761 bp in length and encodes for a protein of 64.3 kDa. Both partial amino acid sequences obtained from GCDA, the N-terminus as well as an internal tryptic peptide, were detected in the open reading frame (ORF) of gcdA. 2. Sequencing of the flanking regions revealed three adjacent ORF (ORF1-3) which do not code for any of the peptide sequences known of the other GCD subunits. The ORF downstream of gcdA (ORF3) is followed by hgdA and hgdB coding for 2-hydroxyglutaryl-CoA dehydratase, the preceding enzyme of the pathway of glutamate fermentation. Our results suggest that at least these three genes of the hydroxyglutarate pathway are organised in an operon and that the genes of the other GCD subunits from which peptide sequences are known (GCDB and GCDC) are not located adjacent to gcdA. 3. gcdA was amplified from genomic DNA using the polymerase chain reaction and cloned into the expression vector pJF118HE. Active GCDA subunit (up to 2.8 nkat/mg protein), catalysing the biotin-dependent formation of crotonyl-CoA from glutaconyl-CoA, was obtained in cell-free extracts of Escherichia coli DH5 alpha by moderately inducing the tac promoter of pJF118HE with 25-100 microM isopropyl-1-thio-beta-D-galactoside. Strong induction (1 mM isopropyl-1-thio-beta-D-galactoside) led to the formation of inclusion bodies from which GCDA could not be reactivated. The apparent Km = 51 mM for free biotin of the expressed GCDA subunit with V = 1.9 nkat/mg protein is similar to that of butanol-treated GCD composed of GCDA and GCDC (apparent Km = 40 mM). Biocytin was found to be a somewhat better carboxy acceptor for the expressed GCDA subunit (apparent Km = 13 mM; V = 1.0 nkat/mg protein). 4. Native GCD and expressed GCDA were treated with 2 mM N-ethylmaleimide showing different kinetics of inactivation: GCD lost half of its activity within 6 min, whereas expressed GCDA required 21 min.
Topics: Amino Acid Sequence; Base Sequence; Biotin; Carboxy-Lyases; Cloning, Molecular; Escherichia coli; Ethylmaleimide; Gene Expression; Molecular Sequence Data; Oligonucleotide Probes; Open Reading Frames; Polymerase Chain Reaction; Recombinant Proteins; Sequence Analysis, DNA; Sodium-Potassium-Exchanging ATPase; Transformation, Bacterial; Veillonellaceae
PubMed: 8382157
DOI: 10.1111/j.1432-1033.1993.tb17598.x -
European Journal of Biochemistry Apr 1986The steric course of the decarboxylation of glutaconyl-CoA to crotonyl-CoA, catalysed by the biotin-dependent sodium pump glutaconyl-CoA decarboxylase from...
The steric course of the decarboxylation of glutaconyl-CoA to crotonyl-CoA, catalysed by the biotin-dependent sodium pump glutaconyl-CoA decarboxylase from Acidaminococcus fermentans, was elucidated using the sequence: chiral acetate----citrate----glutamate----glutaconyl-CoA----crotonyl-CoA ----chiral acetate. Since glutaconyl-CoA or glutaconate labeled at C-4 was subjected to rapid chemical or enzymatic exchanges, glutamate was fermented to acetate by growing cells of A. fermentans. The analysis of the final chiral acetates gave following deviations from 50% in the fumarase exchange: + 13.8% starting with (R)-acetate and - 13.9% starting with (S)-acetate. The results demonstrated a retention of configuration during the decarboxylation. Thus glutaconyl-CoA decarboxylase adds to the list of biotin enzymes in which exclusive retention of configuration was observed. Glutaconate CoA-transferase from A. fermentans catalysed a 3H exchange of [2,4,4-3H]glutaconate with water when acetyl-CoA was present. At low concentration of acetyl-CoA (20 microM) the exchange ceased after exactly one atom 3H was released into the water, at high concentrations (1 mM) the exchange proceeded further. The apparent Km of acetyl-CoA in the exchange (1.1 microM) was 150 times smaller than that of the complete CoA transfer. It was concluded that either a mixed anhydride, between a carboxyl group of the enzyme and [2,4,4-3H]glutaconate, or enzyme-bound glutaconyl-CoA was the exchanging species.
Topics: Acetates; Biotin; Carboxy-Lyases; Catalysis; Fermentation; Ion Channels; Kinetics; Molecular Conformation; Sodium; Substrate Specificity; Veillonellaceae
PubMed: 2422028
DOI: 10.1111/j.1432-1033.1986.tb09576.x -
Biochemistry May 20022-hydroxyglutaryl-CoA dehydratase from Acidaminococcus fermentans catalyzes the chemical difficult elimination of water from (R)-2-hydroxyglutaryl-CoA to glutaconyl-CoA....
2-hydroxyglutaryl-CoA dehydratase from Acidaminococcus fermentans catalyzes the chemical difficult elimination of water from (R)-2-hydroxyglutaryl-CoA to glutaconyl-CoA. The enzyme consists of two oxygen-sensitive protein components, the homodimeric activator (A) with one [4Fe-4S]1+/2+ cluster and the heterodimeric dehydratase (D) with one nonreducible [4Fe-4S]2+ cluster and reduced riboflavin 5'-monophosphate (FMNH2). For activation, ATP, Mg2+, and a reduced flavodoxin (16 kDa) purified from A. fermentans are required. The [4Fe-4S](1+/2+) cluster of component A is exposed to the solvent since it is accessible to iron chelators. Upon exchange of the bound ADP by ATP, the chelation rate is 8-fold enhanced, indicating a large conformational change. Oxidized component A exhibits ATPase activity of 6 s(-1), which is completely abolished upon reduction by one electron. UV-visible spectroscopy revealed a spontaneous one-electron transfer from flavodoxin hydroquinone (E(0)' = -430 mV) to oxidized component A, whereby the [4Fe-4S]2+ cluster of component A became reduced. Combined kinetic, EPR, and Mössbauer spectrocopic investigations exhibited an ATP-dependent oxidation of component A by component D. Whereas the [4Fe-4S]2+ cluster of component D remained in the oxidized state, a new EPR signal became visible attributed to a d1-metal species, probably Mo(V). Metal analysis with neutron activation and atomic absorption spectroscopy gave 0.07-0.2 Mo per component D. In summary, the data suggest that in the presence of ATP one electron is transferred from flavodoxin hydroquinone via the [4Fe-4S]1+/2+ cluster of component A to Mo(VI) of component D, which is thereby reduced to Mo(V). The latter may supply the electron necessary for transient charge reversal in the unusual dehydration.
Topics: Adenosine Triphosphate; Amino Acid Sequence; Clostridium; Electron Spin Resonance Spectroscopy; Electron Transport; Flavodoxin; Hydro-Lyases; Hydrolysis; Sequence Homology, Amino Acid; Spectrophotometry, Ultraviolet
PubMed: 11980491
DOI: 10.1021/bi020033m