Primary alcohols and di-alcohols as growth substrates for the purple nonsulfur bacteriumRhodobacter capsulatus†

2000 ◽  
Vol 46 (12) ◽  
pp. 1166-1170
Author(s):  
Panagiotis E Pantazopoulous ◽  
Michael T Madigan
Keyword(s):  
Fatty Acids ◽  
Organic Acids ◽  
Rhodobacter Capsulatus ◽  
Carbon Sources ◽  
Purple Nonsulfur Bacteria ◽  
Alcohol Metabolism ◽  
Primary Alcohols ◽  
Tertiary Alcohols ◽  
Unsaturated Alcohols ◽  
Growth Experiments

Growth experiments were performed with the purple nonsulfur bacterium Rhodobacter capsulatus to test its ability to use aliphatic, methyl-substituted, and unsaturated alcohols, as well as di-alcohols, as carbon sources for growth. Both phototrophic and chemotrophic growth was observed on a wide variety of such alcohols. By contrast, secondary or tertiary alcohols, or primary alcohols containing an ethyl or propyl substituent, did not support growth. In addition, preculture history and serial subculturing were found to be important factors for obtaining reliable growth of R. capsulatus on alcohols. Collectively, these results suggest that the carbon nutritional diversity of Rhodobacter capsulatus is even greater than previously suspected and that besides metabolizing organic acids and fatty acids in nature, this species may also be a major consumer of alcohols.Key words: purple nonsulfur bacteria, Rhodobacter capsulatus, alcohol metabolism, primary alcohols, di-alcohols.

scholarly journals Photoheterotrophic Assimilation of Valerate and Associated Polyhydroxyalkanoate Production by Rhodospirillum rubrum

2020 ◽  
Vol 86 (18) ◽  
Author(s):  
Guillaume Bayon-Vicente ◽  
Sarah Zarbo ◽  
Adam Deutschbauer ◽  
Ruddy Wattiez ◽  
Baptiste Leroy
Keyword(s):  
Fatty Acids ◽  
Volatile Fatty Acids ◽  
Rhodospirillum Rubrum ◽  
Carbon Sources ◽  
Industrial Applications ◽  
Purple Nonsulfur Bacteria ◽  
Content Type ◽  
Fermentation Processes ◽  
Bicarbonate Ions ◽  
Pha Production

ABSTRACT Purple nonsulfur bacteria are increasingly recognized for industrial applications in bioplastics, pigment, and biomass production. In order to optimize the yield of future biotechnological processes, the assimilation of different carbon sources by Rhodospirillum rubrum has to be understood. As they are released from several fermentation processes, volatile fatty acids (VFAs) represent a promising carbon source in the development of circular industrial applications. To obtain an exhaustive characterization of the photoheterotrophic metabolism of R. rubrum in the presence of valerate, we combined phenotypic, proteomic, and genomic approaches. We obtained evidence that valerate is cleaved into acetyl coenzyme A (acetyl-CoA) and propionyl-CoA and depends on the presence of bicarbonate ions. Genomic and enzyme inhibition data showed that a functional methylmalonyl-CoA pathway is essential. Our proteomic data showed that the photoheterotrophic assimilation of valerate induces an intracellular redox stress which is accompanied by an increased abundance of phasins (the main proteins present in polyhydroxyalkanoate [PHA] granules). Finally, we observed a significant increase in the production of the copolymer P(HB-co-HV), accounting for a very high (>80%) percentage of HV monomer. Moreover, an increase in the PHA content was obtained when bicarbonate ions were progressively added to the medium. The experimental conditions used in this study suggest that the redox imbalance is responsible for PHA production. These findings also reinforce the idea that purple nonsulfur bacteria are suitable for PHA production through a strategy other than the well-known feast-and-famine process. IMPORTANCE The use and the littering of plastics represent major issues that humanity has to face. Polyhydroxyalkanoates (PHAs) are good candidates for the replacement of oil-based plastics, as they exhibit comparable physicochemical properties but are biobased and biodegradable. However, the current industrial production of PHAs is curbed by the production costs, which are mainly linked to the carbon source. Volatile fatty acids issued from the fermentation processes constitute interesting carbon sources, since they are inexpensive and readily available. Among them, valerate is gaining interest regarding the ability of many bacteria to produce a copolymer of PHAs. Here, we describe the photoheterotrophic assimilation of valerate by Rhodospirillum rubrum, a purple nonsulfur bacterium mainly known for its metabolic versatility. Using a knowledge-based optimization process, we present a new strategy for the improvement of PHA production, paving the way for the use of R. rubrum in industrial processes.

scholarly journals Functional analysis of the fatty acid and alcohol metabolism of Pseudomonas putida using RB-TnSeq

2020 ◽  
Author(s):  
Mitchell G. Thompson ◽  
Matthew R. Incha ◽  
Allison N. Pearson ◽  
Matthias Schmidt ◽  
William A. Sharpless ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Metabolic Engineering ◽  
Pseudomonas Putida ◽  
Genetic Basis ◽  
Carbon Sources ◽  
Alcohol Metabolism ◽  
Initial Oxidation ◽  
Genetic Changes ◽  
Substrate Preferences

ABSTRACTWith its ability to catabolize a wide variety of carbon sources and a growing engineering toolkit, Pseudomonas putida KT2440 is emerging as an important chassis organism for metabolic engineering. Despite advances in our understanding of this organism, many gaps remain in our knowledge of the genetic basis of its metabolic capabilities. These gaps are particularly noticeable in our understanding of both fatty acid and alcohol catabolism, where many paralogs putatively coding for similar enzymes co-exist making biochemical assignment via sequence homology difficult. To rapidly assign function to the enzymes responsible for these metabolisms, we leveraged Random Barcode Transposon Sequencing (RB-TnSeq). Global fitness analyses of transposon libraries grown on 13 fatty acids and 10 alcohols produced strong phenotypes for hundreds of genes. Fitness data from mutant pools grown on varying chain length fatty acids indicated specific enzyme substrate preferences, and enabled us to hypothesize that DUF1302/DUF1329 family proteins potentially function as esterases. From the data we also postulate catabolic routes for the two biogasoline molecules isoprenol and isopentanol, which are catabolized via leucine metabolism after initial oxidation and activation with CoA. Because fatty acids and alcohols may serve as both feedstocks or final products of metabolic engineering efforts, the fitness data presented here will help guide future genomic modifications towards higher titers, rates, and yields.IMPORTANCETo engineer novel metabolic pathways into P. putida, a comprehensive understanding of the genetic basis of its versatile metabolism is essential. Here we provide functional evidence for the putative roles of hundreds of genes involved in the fatty acid and alcohol metabolism of this bacterium. These data provide a framework facilitating precise genetic changes to prevent product degradation and channel the flux of specific pathway intermediates as desired.

scholarly journals Fatty Acid and Alcohol Metabolism in Pseudomonas putida: Functional Analysis Using Random Barcode Transposon Sequencing

2020 ◽  
Vol 86 (21) ◽  
Author(s):  
Mitchell G. Thompson ◽  
Matthew R. Incha ◽  
Allison N. Pearson ◽  
Matthias Schmidt ◽  
William A. Sharpless ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Metabolic Engineering ◽  
Pseudomonas Putida ◽  
Genetic Basis ◽  
Carbon Sources ◽  
Alcohol Metabolism ◽  
Initial Oxidation ◽  
Content Type ◽  
Substrate Preferences

ABSTRACT With its ability to catabolize a wide variety of carbon sources and a growing engineering toolkit, Pseudomonas putida KT2440 is emerging as an important chassis organism for metabolic engineering. Despite advances in our understanding of the organism, many gaps remain in our knowledge of the genetic basis of its metabolic capabilities. The gaps are particularly noticeable in our understanding of both fatty acid and alcohol catabolism, where many paralogs putatively coding for similar enzymes coexist, making biochemical assignment via sequence homology difficult. To rapidly assign function to the enzymes responsible for these metabolisms, we leveraged random barcode transposon sequencing (RB–Tn-Seq). Global fitness analyses of transposon libraries grown on 13 fatty acids and 10 alcohols produced strong phenotypes for hundreds of genes. Fitness data from mutant pools grown on fatty acids of varying chain lengths indicated specific enzyme substrate preferences and enabled us to hypothesize that DUF1302/DUF1329 family proteins potentially function as esterases. From the data, we also postulate catabolic routes for the two biogasoline molecules isoprenol and isopentanol, which are catabolized via leucine metabolism after initial oxidation and activation with coenzyme A (CoA). Because fatty acids and alcohols may serve as both feedstocks and final products of metabolic-engineering efforts, the fitness data presented here will help guide future genomic modifications toward higher titers, rates, and yields. IMPORTANCE To engineer novel metabolic pathways into P. putida, a comprehensive understanding of the genetic basis of its versatile metabolism is essential. Here, we provide functional evidence for the putative roles of hundreds of genes involved in the fatty acid and alcohol metabolism of the bacterium. These data provide a framework facilitating precise genetic changes to prevent product degradation and to channel the flux of specific pathway intermediates as desired.

scholarly journals Synthesis of fatty acids in the perfused mouse liver

1974 ◽  
Vol 142 (3) ◽  
pp. 611-618 ◽  
Author(s):  
D. Michael W. Salmon ◽  
Neil L. Bowen ◽  
Douglas A. Hems
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Fatty Acid Synthesis ◽  
De Novo ◽  
Saturated Fatty Acids ◽  
Carbon Sources ◽  
Acid Synthesis ◽  
Hepatic Glycogen ◽  
Total Rate ◽  
Glucose Carbon

1. Fatty acid synthesis de novo was measured in the perfused liver of fed mice. 2. The total rate, measured by the incorporation into fatty acid of3H from3H2O (1–7μmol of fatty acid/h per g of fresh liver), resembled the rate found in the liver of intact mice. 3. Perfusions with l-[U-14C]lactic acid and [U-14C]glucose showed that circulating glucose at concentrations less than about 17mm was not a major carbon source for newly synthesized fatty acid, whereas lactate (10mm) markedly stimulated fatty acid synthesis, and contributed extensive carbon to lipogenesis. 4. The identification of 50% of the carbon converted into newly synthesized fatty acid lends further credibility to the use of3H2O to measure hepatic fatty acid synthesis. 5. The total rate of fatty acid synthesis, and the contribution of glucose carbon to lipogenesis, were directly proportional to the initial hepatic glycogen concentration. 6. The proportion of total newly synthesized lipid that was released into the perfusion medium was 12–16%. 7. The major products of lipogenesis were saturated fatty acids in triglyceride and phospholipid. 8. The rate of cholesterol synthesis, also measured with3H2O, expressed as acetyl residues consumed, was about one-fourth of the basal rate of fatty acid synthesis. 9. These results are discussed in terms of the carbon sources of hepatic newly synthesized fatty acids, and the effect of glucose, glycogen and lactate in stimulating lipogenesis, independently of their role as precursors.

Cloning of the Nocardia corallina polyhydroxyalkanoate synthase gene and production of poly-(3-hydroxybutyrate-co-3-hydroxyhexanoate) and poly-(3-hydroxyvalerate-co-3-hydroxyheptanoate)

1998 ◽  
Vol 44 (7) ◽  
pp. 687-691 ◽  
Author(s):  
Brian Hall ◽  
Jennifer Baldwin ◽  
Ho Gun Rhie ◽  
Douglas Dennis
Keyword(s):  
Fatty Acids ◽  
Ralstonia Eutropha ◽  
Carbon Sources ◽  
Pha Synthase ◽  
Broad Host Range ◽  
Coding Region ◽  
Reading Frame ◽  
Nocardia Corallina ◽  
Odd Chain Fatty Acids ◽  
Chain Fatty Acids

The polyhydroxyalkanoate (PHA) synthase gene (phaCNc) from Nocardia corallina was identified in a lambda library on a 6-kb BamHI fragment. A 2.8-kb XhoII subfragment was found to contain the ntact PHA synthase. This 2.8-kb fragment was subjected to DNA sequencing and was found to contain the coding region for the PHA synthase and a small downstream open reading frame of unknown function. On the basis of DNA sequence, phaCNc is closest in homology to the PHA synthases (phaCPaI and phaCPaII) of Pseudomonas aeruginosa (approximately 41% identity and 55% similarity). The 2.8-kb XhoII fragment containing phaCNc was subcloned into broad host range mobilizable plasmids and transferred into Escherichia coli, Klebsiella aerogenes (both containing a plasmid bearing phaA and phaB from Ralstonia eutropha), and PHA-negative strains of R. eutropha and Pseudomonas putida. The recombinant strains were grown on various carbon sources and the resulting polymers were analyzed. In these strains, the PHA synthase from N. corallina was able to mediate the production of poly(3-hydroxybutyrate-co-3-hydroxy-hexanoate) containing high levels of 3-hydroxyhexanoate when grown on hexanoate and larger even-chain fatty acids and poly(3-hydroxyvalerate-co-3-hydroxyheptanoate) containing high levels of 3-hydroxyheptanoate when grown on heptanoate or larger odd-chain fatty acids. Key words: polyhydroxyalkanoates (PHAs), Nocardia corallina, biodegradable, polyester.

scholarly journals Paucibacter toxinivorans gen. nov., sp. nov., a bacterium that degrades cyclic cyanobacterial hepatotoxins microcystins and nodularin

2005 ◽  
Vol 55 (4) ◽  
pp. 1563-1568 ◽  
Author(s):  
Jarkko Rapala ◽  
Katri A. Berg ◽  
Christina Lyra ◽  
R. Maarit Niemi ◽  
Werner Manz ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Gene Sequence ◽  
Sequence Similarity ◽  
Carbon Sources ◽  
16S Rrna Gene Sequence ◽  
Rrna Gene ◽  
Rrna Gene Sequence

Thirteen bacterial isolates from lake sediment, capable of degrading cyanobacterial hepatotoxins microcystins and nodularin, were characterized by phenotypic, genetic and genomic approaches. Cells of these isolates were Gram-negative, motile by means of a single polar flagellum, oxidase-positive, weakly catalase-positive and rod-shaped. According to phenotypic characteristics (carbon utilization, fatty acid and enzyme activity profiles), the G+C content of the genomic DNA (66·1–68·0 mol%) and 16S rRNA gene sequence analysis (98·9–100 % similarity) the strains formed a single microdiverse genospecies that was most closely related to Roseateles depolymerans (95·7–96·3 % 16S rRNA gene sequence similarity). The isolates assimilated only a few carbon sources. Of the 96 carbon sources tested, Tween 40 was the only one used by all strains. The strains were able to mineralize phosphorus from organic compounds, and they had strong leucine arylamidase and chymotrypsin activities. The cellular fatty acids identified from all strains were C16 : 0 (9·8–19 %) and C17 : 1 ω7c (<1–5·8 %). The other predominant fatty acids comprised three groups: summed feature 3 (<1–2·2 %), which included C14 : 0 3-OH and C16 : 1 iso I, summed feature 4 (54–62 %), which included C16 : 1 ω7c and C15 : 0 iso OH, and summed feature 7 (8·5–28 %), which included ω7c, ω9c and ω12t forms of C18 : 1. A more detailed analysis of two strains indicated that C16 : 1 ω7c was the main fatty acid. The phylogenetic and phenotypic features separating our strains from recognized bacteria support the creation of a novel genus and species, for which the name Paucibacter toxinivorans gen. nov., sp. nov. is proposed. The type strain is 2C20T (=DSM 16998T=HAMBI 2767T=VYH 193597T).

Identification of a DNA region required for growth of Pseudomonas syringae pv. tomato on tomato plants

1992 ◽  
Vol 38 (9) ◽  
pp. 883-890 ◽  
Author(s):  
Dennis P. Jackson ◽  
Douglas A. Gray ◽  
Vincent L. Morris ◽  
Diane A. Cuppels
Keyword(s):  
Organic Acids ◽  
Pseudomonas Syringae ◽  
Acid Metabolism ◽  
Carbon Sources ◽  
Hypersensitive Reaction ◽  
Wild Type ◽  
In Planta ◽  
Tomato Plants ◽  
Tartaric Acids ◽  
Nonpathogenic Strain

The prototrophic Pseudomonas syringae pv. tomato mutant DC3481, which is the result of a single-site Tn5 insertion, cannot grow and cause disease on tomato plants and cannot use the major organic acids of tomato, i.e., citric, malic, succinic, and tartaric acids, as sole carbon sources. Although nonpathogenic, strain DC3481 can still induce a hypersensitive reaction in nonhost plants. We have identified a 30-kb fragment of P. syringae pv. tomato wild-type DNA that can complement this mutant. EcoRI fragments from this region were subcloned and individually subjected to functional complementation analysis. The 3.8-kb fragment, which was the site of the Tn5 insertion, restored pathogenicity and the ability to use all the major organic acids of tomato as carbon sources. It shares sequence homology with several P. syringae pathovars but not other bacterial tomato pathogens. Our results indicate that sequences on the 3.8-kb EcoRI fragment are required for both the ability to grow on tomato leaves (and thus cause disease) and the utilization of carboxylic acids common to tomato. The 3.8-kb fragment may contain a sequence (or sequences) that regulates both traits. Key words: Pseudomonas syringae pv. tomato, phytopathogenicity, Tn5, tricarboxylic acid metabolism, bacterial speck, growth in planta.

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