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D-Gluconic Acid/D-Glucono-δ-lactone Assay Kit

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D-Gluconate Acid D-Glucono-gamma-lactone Assay K-GATE Scheme
   
Product code: K-GATE
€223.00

60 assays (manual) / 600 assays (microplate) / 600 assays (auto-analyser)

Prices exclude VAT

Available for shipping

Content: 60 assays (manual) / 600 assays (microplate) / 600 assays (auto-analyser)
Shipping Temperature: Ambient
Storage Temperature: Short term stability: 2-8oC,
Long term stability: See individual component labels
Stability: > 2 years under recommended storage conditions
Analyte: D-Gluconate, D-Glucono-δ-lactone
Assay Format: Spectrophotometer, Microplate, Auto-analyser
Detection Method: Absorbance
Wavelength (nm): 340
Signal Response: Increase
Linear Range: 0.8 to 50 µg of D-gluconic acid per assay
Limit of Detection: 0.792 mg/L
Reaction Time (min): ~ 6 min
Application examples: Wine, meat, processed meat (e.g. additives), fruit juice, dairy products, pharmaceuticals, paper and other materials (e.g. biological cultures, samples, etc.).
Method recognition: Methods based on this principle have been accepted by ISO, DIN and GOST

The D-Gluconic Acid/D-Glucono-δ-lactone test kit is suitable for the specific measurement and analysis of D-gluconic acid/D-gluconolactone in foods and beverages.

Note for Content: The number of manual tests per kit can be doubled if all volumes are halved.  This can be readily accommodated using the MegaQuantTM  Wave Spectrophotometer (D-MQWAVE).

Check our complete range of monosaccharide assay kits.

Scheme-K-GATE GATE Megazyme

Advantages
  • Extended cofactors stability. Dissolved cofactors stable for > 1 year at 4oC.
  • All reagents stable for > 2 years after preparation 
  • Very competitive price (cost per test) 
  • Very rapid reaction 
  • Mega-Calc™ software tool is available from our website for hassle-free raw data processing 
  • Standard included
  • Suitable for manual, microplate and auto-analyser formats
Documents
Certificate of Analysis
Safety Data Sheet
FAQs Assay Protocol Data Calculator Product Performance Validation Report
Publications
Megazyme publication

Megazyme “advanced” wine test kits general characteristics and validation.

Charnock, S. J., McCleary, B. V., Daverede, C. & Gallant, P. (2006). Reveue des Oenologues, 120, 1-5.

Many of the enzymatic test kits are official methods of prestigious organisations such as the Association of Official Analytical Chemicals (AOAC) and the American Association of Cereal Chemists (AACC) in response to the interest from oenologists. Megazyme decided to use its long history of enzymatic bio-analysis to make a significant contribution to the wine industry, by the development of a range of advanced enzymatic test kits. This task has now been successfully completed through the strategic and comprehensive process of identifying limitations of existing enzymatic bio-analysis test kits where they occurred, and then using advanced techniques, such as molecular biology (photo 1), to rapidly overcome them. Novel test kits have also been developed for analytes of emerging interest to the oenologist, such as yeast available nitrogen (YAN; see pages 2-3 of issue 117 article), or where previously enzymes were simply either not available, or were too expensive to employ, such as for D-mannitol analysis.

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Megazyme publication

Grape and wine analysis: Oenologists to exploit advanced test kits.

Charnock, S. C. & McCleary, B. V. (2005). Revue des Enology, 117, 1-5.

It is without doubt that testing plays a pivotal role throughout the whole of the vinification process. To produce the best possible quality wine and to minimise process problems such as “stuck” fermentation or troublesome infections, it is now recognised that if possible testing should begin prior to harvesting of the grapes and continue through to bottling. Traditional methods of wine analysis are often expensive, time consuming, require either elaborate equipment or specialist expertise and frequently lack accuracy. However, enzymatic bio-analysis enables the accurate measurement of the vast majority of analytes of interest to the wine maker, using just one piece of apparatus, the spectrophotometer (see previous issue No. 116 for a detailed technical review). Grape juice and wine are amenable to enzymatic testing as being liquids they are homogenous, easy to manipulate, and can generally be analysed without any sample preparation.

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Publication

Host factors modulating Ochratoxin A biosynthesis during fruit colonization by Aspergillus carbonarius.

Maor, U., Barda, O., Sadhasivam, S., Bi, Y., Zakin, V., Prusky, D. B. & Sionov, E. (2021). J. Fungi, 7(1), 10.

Aspergillus carbonarius is a strong and consistent ochratoxin A (OTA) producer and considered to be the main source of this toxic metabolite in grapes and grape products such as wine, grape juice and dried vine fruit. OTA is produced under certain growth conditions and its accumulation is affected by several environmental factors, such as growth phase, substrate, temperature, water activity and pH. In this study, we examined the impact of fruit host factors on regulation and accumulation of OTA in colonized grape berries, and assessed in vitro the impact of those factors on the transcriptional levels of the key genes and global regulators contributing to fungal colonization and mycotoxin synthesis. We found that limited sugar content, low pH levels and high malic acid concentrations activated OTA biosynthesis by A. carbonarius, both in synthetic media and during fruit colonization, through modulation of global regulator of secondary metabolism, laeA and OTA gene cluster expression. These findings indicate that fruit host factors may have a significant impact on the capability of A. carbonarius to produce and accumulate OTA in grapes.

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Uncovering a superfamily of nickel-dependent hydroxyacid racemases and epimerases.

Desguin, B., Urdiain-Arraiza, J., Da Costa, M., Fellner, M., Hu, J., Hausinger, R. P., Desmet, T., Hols, P. & Soumillion, P. (2020). Scientific Reports, 10(1), 1-11.

Isomerization reactions are fundamental in biology. Lactate racemase, which isomerizes L- and D-lactate, is composed of the LarA protein and a nickel-containing cofactor, the nickel-pincer nucleotide (NPN). In this study, we show that LarA is part of a superfamily containing many different enzymes. We overexpressed and purified 13 lactate racemase homologs, incorporated the NPN cofactor, and assayed the isomerization of different substrates guided by gene context analysis. We discovered two malate racemases, one phenyllactate racemase, one α-hydroxyglutarate racemase, two D-gluconate 2-epimerases, and one short-chain aliphatic α-hydroxyacid racemase among the tested enzymes. We solved the structure of a malate racemase apoprotein and used it, along with the previously described structures of lactate racemase holoprotein and D-gluconate epimerase apoprotein, to identify key residues involved in substrate binding. This study demonstrates that the NPN cofactor is used by a diverse superfamily of α-hydroxyacid racemases and epimerases, widely expanding the scope of NPN-dependent enzymes.

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The pH-Responsive Transcription Factor PacC Governs Pathogenicity and Ochratoxin A Biosynthesis in Aspergillus carbonarius.

Barda, O., Maor, U., Sadhasivam, S., Bi, Y., Zakin, V., Prusky, D. & Sionov, E. (2020). Frontiers in Microbiology, 11, 210.

Pathogenic fungi must respond effectively to changes in environmental pH for successful host colonization, virulence and toxin production. Aspergillus carbonarius is a mycotoxigenic pathogen with the ability to colonize many plant hosts and secrete ochratoxin A (OTA). In this study, we characterized the functions and addressed the role of PacC-mediated pH signaling in A. carbonarius pathogenicity using designed pacC gene knockout mutant. ΔAcpacC mutant displayed an acidity-mimicking phenotype, which resulted in impaired fungal growth at neutral/alkaline pH, accompanied by reduced sporulation and conidial germination compared to the wild type (WT) strain. The ΔAcpacC mutant was unable to efficiently acidify the growth media as a direct result of diminished gluconic and citric acid production. Furthermore, loss of AcpacC esulted in a complete inhibition of OTA production at pH 7.0. Additionally, ΔAcpacC mutant exhibited attenuated virulence compared to the WT toward grapes and nectarine fruits. Reintroduction of pacC gene into ΔAcpacC mutant restored the WT phenotype. Our results demonstrate important roles of PacC of A. carbonarius in OTA biosynthesis and in pathogenicity by controlling transcription of genes important for fungal secondary metabolism and infection.

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A simple enzymatic assay for the quantification of C1-specific cellulose oxidation by lytic polysaccharide monooxygenases.

Keller, M. B., Felby, C., Labate, C. A., Pellegrini, V. O. A., Higasi, P., Singh, R. K., Polikarpov, I. & Blossom, B. M. (2020). Biotechnology Letters, 42(1), 93-102.

Objective: The development of an enzymatic assay for the specific quantification of the C1-oxidation product, i.e. gluconic acid of cellulose active lytic polysaccharide monooxygenases (LPMOs). Results: In combination with a β-glucosidase, the spectrophotometrical assay can reliably quantify the specific C1-oxidation product of LPMOs acting on cellulose. It is applicable for a pure cellulose model substrate as well as lignocellulosic biomass. The enzymatic assay compares well with the quantification performed by HPAEC-PAD. In addition, we show that simple boiling is not sufficient to inactivate LPMOs and we suggest to apply a metal chelator in addition to boiling or to drastically increase pH for proper inactivation. Conclusions: We conclude that the versatility of this simple enzymatic assay makes it useful in a wide range of experiments in basic and applied LPMO research and without the need for expensive instrumentation, e.g. HPAEC-PAD.

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A 2-year multisite study of viticultural and environmental factors affecting rotundone concentration in Duras red wine.

Geffroy, O., Descôtes, J., Levasseur-Garcia, C., Debord, C., Denux, J. P. & Dufourcq, T. (2019). OENO One, 53(3).

Aim: A study was carried out in 2013 and 2014 to determine the key environmental and viticultural variables affecting the concentration of rotundone, the black pepper aroma compound, in Vitis vinifera L. cv. Duras red wines at 10 different vineyard blocks. Methods and Results: For each block, data for fruit quality attributes, as well as climatic and agronomical variables, were collected. Rotundone was quantified in wines prepared by microvinification techniques (in a 1-L Erlenmeyer flask). Rotundone concentration varied across blocks from 63 ng/L to 239 ng/L in 2013 and from 25 ng/L to 115 ng/L in 2014. Three separate partial least squares regression models were constructed to predict rotundone concentration in wines in 2013, in 2014, and in both vintages. Gluconic acid, a secondary metabolite of Botrytis cinerea, had a substantial contribution to the 2013 and multivintage models, with a negative regression coefficient with rotundone concentration. Other predictors were associated with abiotic factors such as cumulative rainfall, thermal index, hours of sunshine and mean daily irradiation. Conclusions: Our results indicate that mesoscale climatic variables are the key factors determining rotundone concentration, and also suggest that Botrytis cinerea may be involved in rotundone degradation. Significance and impact of the study: Our findings may assist grape growers producing Duras red wines to select specific vineyard blocks with the aim of producing wines with a desired rotundone concentration. They also open up new fields of investigation into mechanisms involved in possible rotundone degradation by Botrytis cinerea.

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Eurypsychrophilic Pseudomonas spp. isolated from Venezuelan tropical glaciers as promoters of wheat growth and biocontrol agents of plant pathogens at low temperatures.

Rondón, J. J., Ball, M. M., Castro, L. T. & Yarzábal, L. A. (2019). Environmental Sustainability, 2(3), 265-275.

Andean tropical glaciers are disappearing rapidly and, consequently, the microbes immured in these frozen environments will be lost forever. Some of these microbes are thought to be potentially useful to develop biotechnological products or processes. Among these microbes, plant-growth promoting (PGP) bacteria have been proposed as valuable tools to develop cold-active biofertilizers and/or biopesticides. A few years ago, we hypothesized that bacteria immured within glacial ice could be effective in promoting plant growth and/or in protecting plants from pathogen infection, at low temperatures. In this study, we aimed at testing some of these traits, with a suitable plant model (Triticum aestivum). In the present study, from a collection of bacteria isolated from Venezuelan tropical glaciers, we selected four Pseudomonas isolates and tested their PGP effects at low temperatures, both in vitro and on wheat plantlets. The isolates grew well over a wide range of low temperatures and were thus classified as eurypsychrophilic. They also displayed well-known PGP traits: solubilization of inorganic phosphates, production of phytohormones and antagonism against a phytopathogenic oomycete (Pythium ultimum). Inoculation of T. aestivum seeds with some of these Pseudomonas spp. isolates promoted a significant elongation of their roots and shoots. This was also the case when wheat plantlets were grown in sterile sand or soil, at 15°C. Inoculation of wheat seeds also protected plantlets against the damage caused by P. ultimum. Together, our results suggest that some of these Pseudomonas spp. isolates could act as cold-active biofertilizers and/or biocontrol agents.

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Specific molecular interactions between vitis vinifera and botrytis cinerea are required for noble rot development in grape berries.

Lovato, A., Zenoni, S., Tornielli, G. B., Colombo, T., Vandelle, E. & Polverari, A. (2019). Postharvest Biology and Technology, 156, 110924.

Under peculiar climatic conditions, the beneficial form of the necrotrophic fungus Botrytis cinerea can develop on grape berries as a latent infection, known as noble rot, which induces positive biochemical and metabolic changes in the berries, including an increase in the sugar content and the production of aromatic compounds that improve wine quality. The infected berries undergo rapid withering, which is required to produce famous sweet white wines such as Sauternes and Tokaj. To gain insight into the molecular interactions between grapevine berries (Vitis vinifera) and B. cinerea during the establishment of noble rot, we prepared a large-scale transcriptomics dataset representing noble rot development and carried out a comparative meta-analysis with gray mold infection and natural post-harvest withering. In particular, we artificially induced berry botrytization of two grape varieties (Garganega and Möller-Thurgau) and sampled them at different stages of noble rot for comparative whole-transcriptome analysis, highlighting important common transcriptional reprogramming in both varieties reflecting an accelerated withering process. Simultaneously, we analyzed the modulation of B. cinerea genes and compared the expression profile during noble rot development with the previously reported gray mold infection profile, revealing the onset of an infection process by the fungus in its beneficial form associated with reduced virulence. This, together with the restrained plant defense response observed in botrytized berries, may favour the development of noble rot instead of gray mold. Finally, the comprehensive meta-analysis of gene expression during noble rot infection, gray mold and post-harvest withering led to the identification of key genes specifically modulated during noble rot infection.

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Effect of ethanol supplementation on the transcriptional landscape of bionanocellulose producer Komagataeibacter xylinus E25.

Ryngajłło, M., Jacek, P., Cielecka, I., Kalinowska, H. & Bielecki, S. (2019). Applied Microbiology and Biotechnology, 103(16), 6673-6688.

Ethanol exerts a strong positive effect on the cellulose yields from the widely exploited microbial producers of the Komagataeibacter genus. Ethanol is postulated to provide an alternative energy source, enabling effective use of glucose for cellulose biosynthesis rather than for energy acquisition. In this paper, we investigate the effect of ethanol supplementation on the global gene expression profile of Komagataeibacter xylinus E25 using RNA sequencing technology (RNA-seq). We demonstrate that when ethanol is present in the culture medium, glucose metabolism is directed towards cellulose production due to the induction of genes related to UDP-glucose formation and the repression of genes involved in glycolysis and acetan biosynthesis. Transcriptional changes in the pathways of cellulose biosynthesis and c-di-GMP metabolism are also described. The transcript level profiles suggest that Schramm-Hestrin medium supplemented with ethanol promotes bacterial growth by inducing protein biosynthesis and iron uptake. We observed downregulation of genes encoding transposases of the IS110 family which may provide one line of evidence explaining the positive effect of ethanol supplementation on the genotypic stability of K. xylinus E25. The results of this study increase knowledge and understanding of the regulatory effects imposed by ethanol on cellulose biosynthesis, providing new opportunities for directed strain improvement, scaled-up bionanocellulose production, and wider industrial exploitation of the Komagataeibacter species as bacterial cellulose producers.

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Strain Serratia sp. S119: A potential biofertilizer for peanut and maize and a model bacterium to study phosphate solubilization mechanisms.

Ludueña, L. M., Anzuay, M. S., Angelini, J. G., McIntosh, M., Becker, A., Rupp, O., Goesmann, A., Blom, J., Fabra, A. & Taurian, T. (2018). Applied Soil Ecology, In Press.

Strain Serratia sp. S119 is a peanut native bacterium with high phosphate solubilizing activity that promotes the growth of peanut and maize in the cultivation area of Córdoba in Argentina. The aims of this study were to obtain and analyze the genome sequence of Serratia sp. S119 to understand the genetic basis of its beneficial properties on plant growth, and to demonstrate phosphate solubilizing ability in early stages of bacterial growth. Results obtained indicated that soluble P and gluconic acid were detected during exponential growth phase in bacterial supernatant. Analysis of the genome sequence of Serratia sp. S119 obtained from this study showed the presence of genes related to several plant growth promoting traits. The genome sequence of this strain is a valuable source of information to study bacterial response to phosphate starvation and to investigate interaction between this bacterium with host plants under nutritional deficient environments.

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An accurate description of Aspergillus niger organic acid batch fermentation through dynamic metabolic modelling.

Upton, D. J., McQueen-Mason, S. J. & Wood, A. J. (2017). Biotechnology for Biofuels, 10(1), 258.

Background: Aspergillus niger fermentation has provided the chief source of industrial citric acid for over 50 years. Traditional strain development of this organism was achieved through random mutagenesis, but advances in genomics have enabled the development of genome-scale metabolic modelling that can be used to make predictive improvements in fermentation performance. The parent citric acid-producing strain of A. niger, ATCC 1015, has been described previously by a genome-scale metabolic model that encapsulates its response to ambient pH. Here, we report the development of a novel double optimisation modelling approach that generates time-dependent citric acid fermentation using dynamic flux balance analysis. Results: The output from this model shows a good match with empirical fermentation data. Our studies suggest that citric acid production commences upon a switch to phosphate-limited growth and this is validated by fitting to empirical data, which confirms the diauxic growth behaviour and the role of phosphate storage as polyphosphate. Conclusions: The calibrated time-course model reflects observed metabolic events and generates reliable in silico data for industrially relevant fermentative time series, and for the behaviour of engineered strains suggesting that our approach can be used as a powerful tool for predictive metabolic engineering.

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Expressing accessory proteins in cellulolytic Yarrowia lipolytica to improve the conversion yield of recalcitrant cellulose.

Guo, Z. P., Duquesne, S., Bozonnet, S., Nicaud, J. M., Marty, A. & O’Donohue, M. J. (2017). Biotechnology for Biofuels, 10(1), 298.

Background: A recently constructed cellulolytic Yarrowia lipolytica is able to grow efficiently on an industrial organosolv cellulose pulp, but shows limited ability to degrade crystalline cellulose. In this work, we have further engineered this strain, adding accessory proteins xylanase II (XYNII), lytic polysaccharide monooxygenase (LPMO), and swollenin (SWO) from Trichoderma reesei in order to enhance the degradation of recalcitrant substrate. Results: The production of EG I was enhanced using a promoter engineering strategy. This provided a new cellulolytic Y. lipolytica strain, which compared to the parent strain, exhibited higher hydrolytic activity on different cellulosic substrates. Furthermore, three accessory proteins, TrXYNII, TrLPMOA and TrSWO, were individually expressed in cellulolytic and non-cellulolytic Y. lipolytica. The amount of rhTrXYNII and rhTrLPMOA secreted by non-cellulolytic Y. lipolytica in YTD medium during batch cultivation in flasks was approximately 62 and 52 mg/L, respectively. The purified rhTrXYNII showed a specific activity of 532 U/mg-protein on beechwood xylan, while rhTrLPMOA exhibited a specific activity of 14.4 U/g-protein when using the Amplex Red/horseradish peroxidase assay. Characterization of rhTrLPMOA revealed that this protein displays broad specificity against β-(1,4)-linked glucans, but is inactive on xylan. Further studies showed that the presence of TrLPMOA synergistically enhanced enzymatic hydrolysis of cellulose by cellulases, while TrSWO1 boosted cellulose hydrolysis only when it was applied before the action of cellulases. The presence of rTrXYNII enhanced enzymatic hydrolysis of an industrial cellulose pulp and of wheat straw. Co-expressing TrXYNII and TrLPMOA in cellulolytic Y. lipolytica with enhanced EG I production procured a novel engineered Y. lipolytica strain that displayed enhanced ability to degrade both amorphous (CIMV-cellulose) and recalcitrant crystalline cellulose in complex biomass (wheat straw) by 16 and 90%, respectively. Conclusions: This study has provided a potent cellulose-degrading Y. lipolytica strain that co-expresses a core set of cellulolytic enzymes and some accessory proteins. Results reveal that the tuning of cellulase production and the production of accessory proteins leads to optimized performance. Accordingly, the beneficial effect of accessory proteins for cellulase-mediated degradation of cellulose is underlined, especially when crystalline cellulose and complex biomass are used as substrates. Findings specifically underline the benefits and specific properties of swollenin. Although in our study swollenin clearly promoted cellulase action, its use requires process redesign to accommodate its specific mode of action.

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Revalorization of strawberry surpluses by bio-transforming its glucose content into gluconic acid.

Cañete-Rodríguez, A. M., Santos-Dueñas, I. M., Jiménez-Hornero, J. E., Torija-Martínez, M. J., Mas, A. & García-García, I. (2016). Food and Bioproducts Processing, 99, 188-196.

Modern societies produce massive surpluses of food, by-products and wastes that increase the interest for their revalorization. This work examines the use of a culture of Gluconobacter japonicus CECT 8443, without pH control, to convert selectively the glucose content of industrially pasteurized strawberry purée into gluconic acid for the development of new beverages. However, depending on the initial concentration of glucose, the microorganism could transform the acid formed into other compounds; for this reason, in this work the effect of initial sugar concentration on the preservation of the acid was investigated. The results show that the gluconic acid formed in strawberry purée containing no added sugars started to disappear after glucose depletion, but the acid concentration remained constant if sugar-enriched purée was used. The use of this industrial substrate resulted in the presence of yeasts and hence in some fructose uptake; however, the fructose consumption was negligible until after 20-30 h. The use of food by-products is an excellent opportunity not only to recover valuable compounds but for the development of new chemical and biotechnological approaches for their revalorization. This strategy should improve regional economies and contribute to a sustainable management of these underexploited resources.

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An approach for estimating the maximum specific growth rate of Gluconobacter japonicus in strawberry purée without cell concentration data.

Cañete-Rodríguez, A. M., Santos-Dueñas, I. M., Jiménez-Hornero, J. E., Torija-Martínez, M. J., Mas, A. & García-García, I. (2016). Biochemical Engineering Journal, 105, 314-320.

The estimation of the maximum specific growth rate (µ max) for non-readily culturable bacteria, growing on complex media containing suspended solids, is a difficult task considering the important problems in obtaining reliable measures of cell concentration. An example of this situation can be a culture of Gluconobacter japonicus growing in strawberry purée for producing gluconic acid. Based on the dependency between energy requirements of the genus Gluconobacter and substrate uptake as well as its constant relationship between gluconic acid production and total substrate uptake, the total substrate concentration profile during the exponential growth phase could be used for estimating µ max without cell concentration measures. In this case, the high selectivity of the strain for glucose in comparison to fructose resulted in no fructose consumption during the batch; so, just using the glucose concentrations data during the exponential phase allow us to obtain an estimation of µ max Additionally, a rough estimation of the apparent and stoichiometric yields of cell on glucose is also possible.

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Applying systems biology tools to study n‐butanol degradation in Pseudomonas putida KT2440.

Vallon, T., Simon, O., Rendgen‐Heugle, B., Frana, S., Mückschel, B., Broicher, A., Siemann-Herzberg, M., Pfannenstiel, J., Hauer, B., Huber, A., Breuer, M. & Breuer, M. (2015). Engineering in Life Sciences, 15(8), 760-771.

To smoothen the process of n-butanol formation in Pseudomonas putida KT2440, detailed knowledge of the impact of this organic solvent on cell physiology and regulation is of outmost importance. Here, we conducted a detailed systems biology study to elucidate cellular responses at the metabolic, proteomic, and transcriptional level. Pseudomonas putida KT2440 was cultivated in multiple chemostat fermentations using n-butanol either as sole carbon source or together with glucose. Pseudomonas putida KT2440 revealed maximum growth rates (µ) of 0.3 h-1 with n-butanol as sole carbon source and of 0.4 h-1 using equal C-molar amounts of glucose and n-butanol. While C-mole specific substrate consumption and biomass/substrate yields appeared equal at these growth conditions, the cellular physiology was found to be substantially different: adenylate energy charge levels of 0.85 were found when n-butanol served as sole carbon source (similar to glucose as sole carbon source), but were reduced to 0.4 when n-butanol was coconsumed at stable growth conditions. Furthermore, characteristic maintenance parameters changed with increasing n-butanol consumption. 13C flux analysis revealed that central metabolism was split into a glucose-fueled Entner–Doudoroff/pentose-phosphate pathway and an n-butanol-fueled tricarboxylic acid cycle when both substrates were coconsumed. With the help of transcriptome and proteome analysis, the degradation pathway of n-butanol could be unraveled, thus representing an important basis for rendering P. putida KT2440 from an n-butanol consumer to a producer in future metabolic engineering studies.

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Rapid Assessment of Gray Mold (Botrytis cinerea) Infection in Grapes Using Biosensors System.

Cinquanta, L., Albanese, D., De Curtis, F., Malvano, F., Crescitelli, A. & Di Matteo, M. (2015). American Journal of Enology and Viticulture, ajev-2015.

Botrytis cinerea is responsible for the gray mold disease, which causes considerable economic losses for winemakers. Its evaluation in wine grapes is commonly performed through visual estimation, which was demonstrated to be prone to assessor bias. Rapid and simple enzymatic carbon screen printed amperometric biosensors were here used to evaluate gluconic acid and glycerol content on wine grapes at different B. cinerea infection degrees. The lower concentrations measurable by screen-printed amperometric biosensors were 3 mg/L for gluconic acid (corresponding to an infection degree lower than 1%) and 35 mg/L for glycerol; the response times with a flow rate of 0.5 mL/min were in a range of 0.5 to 2 min in the linear ranges. This study demonstrates the effectiveness of the biosensors for rapid analysis of gluconic acid and glycerol in grapes, confirming their high correlation with B. cinerea degree of infection (R2 = 0.98). Thus, the biosensor developed to measure gluconic acid in grapes (or must), was more precise, and gave a faster response than methods that currently exist allowing the percentage of infection of grape berries by B. cinerea to be evaluated.

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Genetic diversity of phosphate-solubilizing peanut (Arachis hypogaea L.) associated bacteria and mechanisms involved in this ability.

Anzuay, M. S., Frola, O., Angelini, J. G., Ludueña, L. M., Fabra, A. & Taurian, T. (2013). Symbiosis , 60(3), 143-154.

In this study, attempts were made to analyze mechanisms involved in the bacterial phosphate-solubilizing ability of peanut isolates. Bacteria were taxonomically identified by analysis of 16S rDNA sequence. Levels of soluble P released by the isolates in unbuffered or buffered with Tris–HCl or MES NBRIP-BPB medium as well as the production of D-gluconic acid were determined in their culture. Presence of two of the genes encoding the cofactor PQQ of GDH enzyme was analyzed in the genome of this bacterial collection. 16S rDNA sequence analysis indicated that isolates belong to genera Serratia, Enterobacter, Pantoea, Acinetobacter, Bacillus and Enterococcus. All bacteria showed ability to solubilize tricalcium phosphate either in unbuffered or buffered medium. Nevertheless, addition of buffer solutions reduced levels of Pi liberated by the isolates. Although almost all isolates produced detectable amounts of D-gluconic acid, no correlation with levels of P soluble released were observed. The presence of pqqE and pqqC genes was detected only in Gram negative bacteria. It was concluded from this study that the mechanism involved in phosphate solubilization is organic acids production and, presence of pqq genes in all Gram negative bacteria analyzed encourages to confirm their role in bacterial phosphate solubilizing ability as well to identify genes involved in this PGP trait in Gram positive bacteria.

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Aerobic deconstruction of cellulosic biomass by an insect-associated Streptomyces.

Takasuka, T. E., Book, A. J., Lewin, G. R., Currie, C. R. & Fox, B. G. (2013). Scientific Reports, 3.

Streptomyces are best known for producing antimicrobial secondary metabolites, but they are also recognized for their contributions to biomass utilization. Despite their importance to carbon cycling in terrestrial ecosystems, our understanding of the cellulolytic ability of Streptomyces is currently limited to a few soil-isolates. Here, we demonstrate the biomass-deconstructing capability of Streptomyces sp. SirexAA-E (ActE), an aerobic bacterium associated with the invasive pine-boring woodwasp Sirex noctilio. When grown on plant biomass, ActE secretes a suite of enzymes including endo- and exo-cellulases, CBM33 polysaccharide-monooxygenases, and hemicellulases. Genome-wide transcriptomic and proteomic analyses, and biochemical assays have revealed the key enzymes used to deconstruct crystalline cellulose, other pure polysaccharides, and biomass. The mixture of enzymes obtained from growth on biomass has biomass-degrading activity comparable to a cellulolytic enzyme cocktail from the fungus Trichoderma reesei, and thus provides a compelling example of high cellulolytic capacity in an aerobic bacterium.

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Serratia marcescens quinoprotein glucose dehydrogenase activity mediates medium acidification and inhibition of prodigiosin production by glucose.

Fender, J. E., Bender, C. M., Stella, N. A., Lahr, R. M., Kalivoda, E. J. & Shanks, R. M. (2012). Applied and Environmental Microbiology, 78(17), 6225-6235.

Serratia marcescens is a model organism for the study of secondary metabolites. The biologically active pigment prodigiosin (2-methyl-3-pentyl-6-methoxyprodiginine), like many other secondary metabolites, is inhibited by growth in glucose-rich medium. Whereas previous studies indicated that this inhibitory effect was pH dependent and did not require cyclic AMP (cAMP), there is no information on the genes involved in mediating this phenomenon. Here we used transposon mutagenesis to identify genes involved in the inhibition of prodigiosin by glucose. Multiple genetic loci involved in quinoprotein glucose dehydrogenase (GDH) activity were found to be required for glucose inhibition of prodigiosin production, including pyrroloquinoline quinone and ubiquinone biosynthetic genes. Upon assessing whether the enzymatic products of GDH activity were involved in the inhibitory effect, we observed that D-glucono-1,5-lactone and D-gluconic acid, but not D-gluconate, were able to inhibit prodigiosin production. These data support a model in which the oxidation of D-glucose by quinoprotein GDH initiates a reduction in pH that inhibits prodigiosin production through transcriptional control of the prodigiosin biosynthetic operon, providing new insight into the genetic pathways that control prodigiosin production. Strains generated in this report may be useful in large-scale production of secondary metabolites.

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Safety Information
Symbol : GHS07
Signal Word : Warning
Hazard Statements : H319
Precautionary Statements : P264, P280, P305+P351+P338, P337+P313
Safety Data Sheet
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