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C-type cytochromes and manganese oxidation in Pseudomonas putida MnB1.

ABSTRACT: Pseudomonas putida MnB1 is an isolate from an Mn oxide-encrusted pipeline that can oxidize Mn(II) to Mn oxides. We used transposon mutagenesis to construct mutants of strain MnB1 that are unable to oxidize manganese, and we characterized some of these mutants. The mutants were divided into three groups: mutants defective in the biogenesis of c-type cytochromes, mutants defective in genes that encode key enzymes of the tricarboxylic acid cycle, and mutants defective in the biosynthesis of tryptophan. The mutants in the first two groups were cytochrome c oxidase negative and did not contain c-type cytochromes. Mn(II) oxidation capability could be recovered in a c-type cytochrome biogenesis-defective mutant by complementation of the mutation.

SUBMITTER: Caspi R

PROVIDER: S-EPMC106463 | biostudies-literature | 1998 Oct

REPOSITORIES: biostudies-literature

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c-type cytochromes and manganese oxidation in Pseudomonas putida MnB1.

Caspi R R Tebo B M BM Haygood M G MG

Applied and environmental microbiology 19981001 10

Pseudomonas putida MnB1 is an isolate from an Mn oxide-encrusted pipeline that can oxidize Mn(II) to Mn oxides. We used transposon mutagenesis to construct mutants of strain MnB1 that are unable to oxidize manganese, and we characterized some of these mutants. The mutants were divided into three groups: mutants defective in the biogenesis of c-type cytochromes, mutants defective in genes that encode key enzymes of the tricarboxylic acid cycle, and mutants defective in the biosynthesis of tryptop ...[more]

PMID: 9758766

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Project description:Pseudomonas putida KT2440 is a well-known model organism for the medium-chain-length (mcl) polyhydroxyalkanoate (PHA) accumulation. (R)-Specific enoyl-coenzyme A hydratase (PhaJ) was considered to be the main supplier of monomers for PHA synthesis by converting the β-oxidation intermediate, trans-2-enoyl-CoA to (R)-3-hydroxyacyl-CoA when fatty acids (FA) are used. Three PhaJ homologues, PhaJ1, PhaJ4 and MaoC, are annotated in P. putida KT2440. To investigate the relationship of fatty acids-PHA metabolism and the role of each PhaJ in PHA biosynthesis in P. putida KT2440, a series of P. putida KT2440 knockouts was obtained. PHA content and monomer composition in wild type (WT) and mutants under different growth conditions were analysed. PhaJ4 was the main monomer supplier for PHA synthesis with FA as sole carbon and energy source, with preference towards C8 and C10 substrate, whereas PhaJ1 showed preference for the C6 substrate. However, when all three PhaJ homologues were deleted, the mutant still accumulated PHA up to 10.7% of the cell dry weight (CDW). The deletion of (R)-3-hydroxydecanoyl-ACP:CoA transacylase (PhaG), which connects de novo FA and PHA synthesis pathways, while causing a further 1.8-fold decrease in PHA content, did not abolish PHA accumulation. Further proteome analysis revealed quinoprotein alcohol dehydrogenases PedE and PedH as potential monomer suppliers, but when these were deleted, the PHA level remained at 2.2-14.8% CDW depending on the fatty acid used and whether nitrogen limitation was applied. Therefore, it is likely that some other non-specific dehydrogenases supply monomers for PHA synthesis, demonstrating the redundancy of PHA metabolism. KEY POINTS: • β-oxidation intermediates are converted to PHA monomers by hydratases PhaJ1, PhaJ4 and MaoC in Pseudomonas putida KT2440. • When these are deleted, the PHA level decreases, but it is not abolished. • PHA non-specific enzyme(s) also contributes to PHA metabolism in KT2440.

Project description:Pseudomonas putida GB-1 is a Mn(II)-oxidizing bacterium that produces pyoverdine-type siderophores (PVDs), which facilitate the uptake of Fe(III) but also influence MnO2 formation. Recently, a non-ribosomal peptide synthetase mutant that does not synthesize PVD was described. Here we identified a gene encoding the PVDGB-1 (PVD produced by strain GB-1) uptake receptor (PputGB1_4082) of strain GB-1 and confirmed its function by in-frame mutagenesis. Growth and other physiological responses of these two mutants and of wild type were compared during cultivation in the presence of three chemically distinct sets of PVDs (siderotypes n°1, n°2, and n°4) derived from various pseudomonads. Under iron-limiting conditions, Fe(III) complexes of various siderotype n°1 PVDs (including PVDGB-1) allowed growth of wild type and the synthetase mutant, but not the receptor mutant, confirming that iron uptake with any tested siderotype n°1 PVD depended on PputGB1_4082. Fe(III) complexes of a siderotype n°2 PVD were not utilized by any strain and strongly induced PVD synthesis. In contrast, Fe(III) complexes of siderotype n°4 PVDs promoted the growth of all three strains and did not induce PVD synthesis by the wild type, implying these complexes were utilized for iron uptake independent of PputGB1_4082. These differing properties of the three PVD types provided a way to differentiate between effects on MnO2 formation that resulted from iron limitation and others that required participation of the PVDGB-1 receptor. Specifically, MnO2 production was inhibited by siderotype n°1 but not n°4 PVDs indicating PVD synthesis or PputGB1_4082 involvement rather than iron-limitation caused the inhibition. In contrast, iron limitation was sufficient to explain the inhibition of Mn(II) oxidation by siderotype n°2 PVDs. Collectively, our results provide insight into how competition for iron via siderophores influences growth, iron nutrition and MnO2 formation in more complex environmental systems.

Dataset Information

C-type cytochromes and manganese oxidation in Pseudomonas putida MnB1.

Publications

c-type cytochromes and manganese oxidation in Pseudomonas putida MnB1.

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