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The central metabolic model for Geobacter sulfurreducens included a single pathway for the biosynthesis of isoleucine that was analogous to that of Escherichia coil, in which the isoleucine precursor 2-oxobutanoate is generated from threonine. [sup.13]C labeling studies performed in G. sulfurreducens indicated that this pathway accounted for a minor fraction of isoleucine biosynthesis and that the majority of isoleucine was instead derived from acetyl-coenzyme A and pyruvate, possibly via the citramalate pathway. Genes encoding citramalate synthase (GSU1798), which catalyzes the first dedicated step in the citramalate pathway, and threonine ammonia-lyase (GSU0486), which catalyzes the conversion of threonine to 2-oxobutanoate, were identified and knocked out. Mutants lacking both of these enzymes were auxotrophs for isoleucine, whereas single mutants were capable of growth in the absence of isoleucine. Biochemical characterization of the single mutants revealed deficiencies in citramalate synthase and threonine ammonia-lyase activity. Thus, in G. sulfurreducens, 2-oxobutanoate can be synthesized either from citramalate or threonine, with the former being the main pathway for isoleucine biosynthesis. The citramalate synthase of G. suifurreducens constitutes the first characterized member of a phylogenetically distinct clade of citramalate synthases, which contains representatives from a wide variety of microorganisms.

Plasmids that facilitate the controlled expression of ppk and ppx genes were constructed and expressed in an Escherichia coli strain to determine the genetic mechanisms of polyphosphate synthesis and degradation. Polyphosphate extraction, enzyme assays and RNA analysis indicated the ability of ppk- and ppx-expressing Escherichia coli strain to produce active polyphosphatekinase (PPK) after the induction of the ppx gene. Furthermore, the growth of phosphate-starved cells was promoted by the phosphate that was released from PPK due to the down-regulation of the phosphate-starvation response.

The article focuses on large-scale sampling of patient microbiomes is giving one organization the tools to correlate microbiome composition with immune health, driving the deployment of safe, live bio therapeutic treatments for cancer and COVID-19. Topics include the role of gut microbes in human health has been recognized for decades, and the recent advances in technologies like next-generation sequencing, transcriptomic and proteomic analyses.

The putative carotenoid biosynthesis pathway in the pink-pigmented facultative methylotroph Methylobacterium extorquens AMI was reconstructed by genomic searches. A colorless mutant was obtained by transposon mutagenesis and the insertion was shown to be in one of the putative phytoene desturase genes.

A mathematical model was developed to describe the physiological co-regulation of two Pseudomonas σ54-dependent promoter/regulator systems, Pu/XylR and Po/DmpR of Pseudomonas strains mt2 and CF600, respectively. Five ordinary differential equations and six algebraic equations were developed to describe the following processes of transcription initiation: binding of the activator protein to the upstream activating sequence, union of the sigma factor with the core polymerase, formation of the open complex, and escape of the transcription machinery from the promoter region. In addition, growth-phase control of the integration host factor (IHF), sigma-70 regulation during stationary phase, and the contribution of (p)ppGpp to both sigma factor selectivity and promoter escape were hypothesized. By including any three of these four effects, the model predicted that expression from both promoters is repressed during exponential growth and sharply increases as the cells enter stationary phase. The difference in behavior of the two systems during overexpression of either σ54 or (p)ppGpp could be explained by different values of two model parameters. To accurately represent the behavior of both promoters in (p)ppGpp null strains, an additional parameter must be varied. Although numerical data available for this system is scarce, the model has proved useful for helping to interpret the experimental observations and to evaluate four hypotheses that have been proposed to explain the phenomenon of exponential silencing. [Copyright &y& Elsevier]

The growth of Methylobacterium extorquens AM1 on C[sub 1] compounds has been well-studied, but little is known about how this methylotroph grows on multicarbon compounds. A Tn5 transposon mutagenesis procedure was performed to identify genes involved in the growth of M. extorquens AM1 on succinate and pyruvate. Of the 15 000 insertion colonies screened, 71 mutants were found that grew on methanol but either grew slowly or were unable to grow on one or both of the multicarbon substrates. For each of these mutants, the chromosomal region adjacent to the insertion site was sequenced, and 55 different genes were identified and assigned putative functions. These genes fell into a number of predicted categories, including central carbon metabolism, carbohydrate metabolism, regulation, transport and non-essential housekeeping functions. This study focused on genes predicted to encode enzymes of central heterotrophic metabolism: 2-oxoglutarate dehydrogenase, pyruvate dehydrogenase and NADH: ubiquinone oxidoreductase. In each case, the mutants showed normal growth on methanol and impaired growth on pyruvate and succinate, consistent with a role specific to heterotrophic metabolism. For the first two cases, no detectable activity of the corresponding enzyme was found in the mutant, verifying the predictions. The results of this study were used to reconstruct multicarbon metabolism of M. extorquens AM1 during growth on methanol, succinate and pyruvate. [ABSTRACT FROM AUTHOR]