Purification and properties of L-alanine dehydrogenase of the phototrophic bacterium Rhodobacter capsulatus E1F1

Caballero, Francisco J.; Cárdenas, Jacobo; Castillo, Francisco

doi:10.1128/jb.171.6.3205-3210.1989

Cited by 37 publications

(19 citation statements)

References 34 publications

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“…3) (28, 29). The amination of pyruvate could be a possible mechanism of ammonia storage or alleviation of ammonia toxicity or could also represent another primary nitrogen assimilation process as in AOB and other bacteria (66,67).…”

Section: Resultsmentioning

confidence: 99%

Proteomics and comparative genomics of Nitrososphaera viennensis reveal the core genome and adaptations of archaeal ammonia oxidizers

Kerou

Offre

Valledor

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

171

196

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Ammonia-oxidizing archaea (AOA) are among the most abundant microorganisms and key players in the global nitrogen and carbon cycles. They share a common energy metabolism but represent a heterogeneous group with respect to their environmental distribution and adaptions, growth requirements, and genome contents. We report here the genome and proteome of Nitrososphaera viennensis EN76, the type species of the archaeal class Nitrososphaeria of the phylum Thaumarchaeota encompassing all known AOA. N. viennensis is a soil organism with a 2.52-Mb genome and 3,123 predicted proteincoding genes. Proteomic analysis revealed that nearly 50% of the predicted genes were translated under standard laboratory growth conditions. Comparison with genomes of closely related species of the predominantly terrestrial Nitrososphaerales as well as the more streamlined marine Nitrosopumilales [Candidatus (Ca.) order] and the acidophile "Ca. Nitrosotalea devanaterra" revealed a core genome of AOA comprising 860 genes, which allowed for the reconstruction of central metabolic pathways common to all known AOA and expressed in the N. viennensis and "Ca. Nitrosopelagicus brevis" proteomes. Concomitantly, we were able to identify candidate proteins for as yet unidentified crucial steps in central metabolisms. In addition to unraveling aspects of core AOA metabolism, we identified specific metabolic innovations associated with the Nitrososphaerales mediating growth and survival in the soil milieu, including the capacity for biofilm formation, cell surface modifications and cell adhesion, and carbohydrate conversions as well as detoxification of aromatic compounds and drugs. ammonia oxidation | proteomics | archaea | comparative genomics | biofilm

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Section: Resultsmentioning

confidence: 99%

Proteomics and comparative genomics of Nitrososphaera viennensis reveal the core genome and adaptations of archaeal ammonia oxidizers

Kerou

Offre

Valledor

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

171

196

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“…Glutamate dehydrogenase provides a lower-affinity non-ATPrequiring system, but this activity was not detected in M. tuberculosis (50). Under certain conditions, alanine dehydrogenase may fulfill an ammonium assimilation role, as has been proposed for other bacteria (6,46). This could be important if glutamine synthetase is inhibited by high levels of alanine or glycine (9).…”

Section: Discussionmentioning

confidence: 97%

ald of Mycobacterium tuberculosis Encodes both the Alanine Dehydrogenase and the Putative Glycine Dehydrogenase

Giffin

Modesti

Raab

et al. 2011

Journal of Bacteriology

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The putative glycine dehydrogenase of Mycobacterium tuberculosis catalyzes the reductive amination of glyoxylate to glycine but not the reverse reaction. The enzyme was purified and identified as the previously characterized alanine dehydrogenase. The Ald enzyme was expressed in Escherichia coli and had both pyruvate and glyoxylate aminating activities. The gene, ald, was inactivated in M. tuberculosis, which resulted in the loss of all activities. Both enzyme activities were found associated with the cell and were not detected in the extracellular filtrate. By using an anti-Ald antibody, the protein was localized to the cell membrane, with a smaller fraction in the cytosol. None was detected in the extracellular medium. The ald knockout strain grew without alanine or glycine and was able to utilize glycine but not alanine as a nitrogen source. Transcription of ald was induced when alanine was the sole nitrogen source, and higher levels of Ald enzyme were measured. Ald is proposed to have several functions, including ammonium incorporation and alanine breakdown. Mycobacterium tuberculosis is the causative agent of tuberculosis, and one of the most successful human pathogens. It was responsible for approximately 2 million deaths in 2008, while currently almost one-third of the world's population is infected with this organism. Research with M. tuberculosis has described a pathogen uniquely adapted to the wide range of harsh environments presented by the host. Much of this work has focused on the microbe's metabolism, with the idea of identification of novel enzymes or pathways to target for drug development.One of these environmental factors is nitrogen availability. Very little is known about the nitrogen sources used by M. tuberculosis in vivo. M. tuberculosis can utilize many amino acids for nitrogen, including alanine and glycine (29). Mutants of M. tuberculosis unable to synthesize proline retained the ability to replicate in the human macrophage cell line THP-1 (35), while other amino acid auxotroph mutants were attenuated (3,17,22,52). A Mycobacterium bovis BCG mutant unable to make methionine showed survival in mice similar to the wild-type strain (32). This suggests some amino acids are available in vivo and serve as nutrients for M. tuberculosis.The enzyme glycine dehydrogenase was first described in M. tuberculosis in 1962 (16). This enzyme was detected by the reductive amination of glyoxylate to glycine concurrent with the oxidation of NADH to NAD ϩ (Fig. 1). This reaction represents glyoxylate reductive aminase (GxRA) activity. The activity corresponding to the reverse reaction, catalyzed by glycine dehydrogenase (GDH), was not detected. The expression of glyoxylate reductive amination by a putative glycine dehydrogenase in M. tuberculosis has been characterized in nonreplicating persistent (NRP) cultures (58). In the Wayne model of dormancy, sealed cultures of M. tuberculosis create a microaerobic environment (NRP-1), which subsequently develops into the anaerobic stage (NRP-2) (58). GxRA activity w...

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“…In Bacillus sp., the main function of alanine dehydrogenase seems to be the generation of pyruvate as an energy source during sporulation (Siranosian et al 1993). When operating in the reverse direction -the reductive amination of pyruvate -the enzyme can be involved in nitrogen assimilation as in, for example, Rhodobacter capsulatus (Caballero et al 1989) 1 Initial steps of the degradation of taurine in Bilophila wadsworthia. The first step is a pyruvate-dependent transamination of taurine, yielding sulfoacetaldehyde and alanine catalysed by taurine:pyruvate aminotransferase.…”

Section: Introductionmentioning

confidence: 99%

Purification, properties and primary structure of alanine dehydrogenase involved in taurine metabolism in the anaerobe Bilophila wadsworthia

Laue

Cook

2000

Archives of Microbiology

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Alanine dehydrogenase [L-alanine:NAD + oxidoreductase (deaminating), EC 1.4.1.4.] catalyses the reversible oxidative deamination of L-alanine to pyruvate and, in the anaerobic bacterium Bilophila wadsworthia RZATAU, it is involved in the degradation of taurine (2-aminoethanesulfonate). The enzyme regenerates the amino-group acceptor pyruvate, which is consumed during the transamination of taurine and liberates ammonia, which is one of the degradation end products. Alanine dehydrogenase seems to be induced during growth with taurine. The enzyme was purified about 24-fold to apparent homogeneity in a three-step purification. SDS-PAGE revealed a single protein band with a molecular mass of 42 kDa. The apparent molecular mass of the native enzyme was 273 kDa, as determined by gel filtration chromatography, suggesting a homo-hexameric structure. The N-terminal amino acid sequence was determined. The pH optimum was pH 9.0 for reductive amination of pyruvate and pH 9.0-11.5 for oxidative deamination of alanine. The apparent K m values for alanine, NAD + , pyruvate, ammonia and NADH were 1.6, 0.15, 1.1, 31 and 0.04 mM, respectively. The alanine dehydrogenase gene was sequenced. The deduced amino acid sequence corresponded to a size of 39.9 kDa and was very similar to that of the alanine dehydrogenase from Bacillus subtilis.

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Purification and properties of L-alanine dehydrogenase of the phototrophic bacterium Rhodobacter capsulatus E1F1

Cited by 37 publications

References 34 publications

Proteomics and comparative genomics of Nitrososphaera viennensis reveal the core genome and adaptations of archaeal ammonia oxidizers

Proteomics and comparative genomics of Nitrososphaera viennensis reveal the core genome and adaptations of archaeal ammonia oxidizers

ald of Mycobacterium tuberculosis Encodes both the Alanine Dehydrogenase and the Putative Glycine Dehydrogenase

Purification, properties and primary structure of alanine dehydrogenase involved in taurine metabolism in the anaerobe Bilophila wadsworthia

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