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Strains derived from HfrH carrying the arcA2 null mutation exhibit a higher respiratory rate, enhanced glucose consumption, and a more-reduced intracellular redox state than arcA deletion mutants of a different lineage. The phenotype of the arcA2 mutants was due to the presence of a creC constitutive mutation introduced by P1 transduction.

The effect of PhaP, a phasin, on bacterial growth and the accumulation of poly(3-hydroxybutyrate) (PHB) from glycerol in bioreactor cultures of recombinant Escherichia coli carrying phaBAC from Azotobacter sp. strain FA8 are examined. The behavior of recombinant Escherichia coli strains carrying phasin genes in a bioreactor is useful in the design of bacterial strains sufficient for the efficient production of polyhydroxyalkanoates (PHAs) from glycerol.

Researchers have isolated the genes responsible for poly(3-hydroxybutyrate) (PHB) synthesis in Azotobacter sp. strain FA8. The genes code for PHB polymerase, beta-ketothiolase and acetoacetyl-coenzyme A reductase. The PHB polymerase can only produce PHB from glucose or octanoate, but not from medium-chain-length hydroxyalkanoates.

Glycerol is used as a substrate for analyzing the effect of eliminating D-lactate synthesis in poly(3-hydroxybutyrate) (PHB)-accumulating recombinant Escherichia coli (K24K). Cofactor measurements have shown that cultures of K24K and K24L have a high intracellular NADPH content and the ldhA mutation has affected cofactor distribution, resulting in a more reduced intracellular state.

Aeromonas salmonicida subsp. pectinolytica 34melT can be considered an extremophile due to the characteristics of the heavily polluted river from which it was isolated. While four subspecies of A. salmonicida are known fish pathogens, 34melT belongs to the only subspecies isolated solely from the environment. Genome analysis revealed a high metabolic versatility, the capability to cope with diverse stress agents, and the lack of several virulence factors found in pathogenic Aeromonas. The most relevant phenotypic characteristics of 34melT are pectin degradation, a distinctive trait of A. salmonicida subsp. pectinolytica, and melanin production. Genes coding for three pectate lyases were detected in a cluster, unique to this microorganism, that contains all genes needed for pectin degradation. Melanin synthesis in 34melT is hypothesized to occur through the homogentisate pathway, as no tyrosinases or laccases were detected and the homogentisate 1,2-dioxygenase gene is inactivated by a transposon insertion, leading to the accumulation of the melanin precursor homogentisate. Comparative genome analysis of other melanogenic Aeromonas strains revealed that this gene was inactivated by transposon insertions or point mutations, indicating that melanin biosynthesis in Aeromonas occurs through the homogentisate pathway. Horizontal gene transfer could have contributed to the adaptation of 34melT to a highly polluted environment, as 13 genomic islands were identified in its genome, some of them containing genes coding for fitness-related traits. Heavy metal resistance genes were also found, along with others associated with oxidative and nitrosative stresses. These characteristics, together with melanin production and the ability to use different substrates, may explain the ability of this microorganism to live in an extremely polluted environment. [ABSTRACT FROM AUTHOR]

Phasins (PhaP) are proteins normally associated with granules of poly(3-hydroxybutyrate) (PHB), a biodegradable polymer accumulated by many bacteria as a reserve molecule. These proteins enhance growth and polymer production in natural and recombinant PHB producers. It has been shown that the production of PHB causes stress in recombinant Escherichia coli, revealed by an increase in the concentrations of several heat stress proteins. In this work, quantitative reverse transcription (qRT)-PCR analysis was used to study the effect of PHB accumulation, and that of PhaP from Azotobacter sp. strain FA8, on the expression of stress-related genes in PHB-producing E. coli. While PHB accumulation was found to increase the transcription of dnaK and ibpA, the expression of these genes and of groES, groEL, rpoH, dps, and yfiD was reduced, when PhaP was coexpressed, to levels even lower than those detected in the non-PHB-accumulating control. These results demonstrated the protective role of PhaP in PHB-synthesizing E. coli and linked the effects of the protein to the expression of stress-related genes, especially ibpA. The effect of PhaP was also analyzed in non-PHB-synthesizing strains, showing that expression of this heterologous protein has an unexpected protective effect in E. coli, under both normal and stress conditions, resulting in increased growth and higher resistance to both heat shock and superoxide stress by paraquat. In addition, PhaP expression was shown to reduce RpoH protein levels during heat shock, probably by reducing or titrating the levels of misfolded proteins. [ABSTRACT FROM AUTHOR]

The CreBC (carbon source-responsive) two-component regulation system of Escherichia coli affects a number of functions, including intermediary carbon catabolism. The impacts of different creC mutations (a ΔcreC mutant and a mutant carrying the constitutive creC510 allele) on bacterial physiology were analyzed in glucose cultures under three oxygen availability conditions. Differences in the amounts of extracellular metabolites produced were observed in the null mutant compared to the wild-type strain and the mutant carrying creC510 and shown to be affected by oxygen availability. The ΔcreC strain secreted more formate, succinate, and acetate but less lactate under low aeration. These metabolic changes were associated with differences in AckA and LdhA activities, both of which were affected by CreC. Measurement of the NAD(P)H/NAD(P)(+) ratios showed that the creC510 strain had a more reduced intracellular redox state, while the opposite was observed for the ΔcreC mutant, particularly under intermediate oxygen availability conditions, indicating that CreC affects redox balance. The null mutant formed more succinate than the wild-type strain under both low aeration and no aeration. Overexpression of the genes encoding phosphoenolpyruvate carboxylase from E. coli and a NADH-forming formate dehydrogenase from Candida boidinii in the ΔcreC mutant further increased the yield of succinate on glucose. Interestingly, the elimination of ackA and adhE did not significantly improve the production of succinate. The diverse metabolic effects of this regulator on the central biochemical network of E. coli make it a good candidate for metabolic-engineering manipulations to enhance the formation of bioproducts, such as succinate., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)