Very high gravity sucrose fermentation by Brazilian industrial yeast strains: effect of nitrogen supplementation

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Evaluation of Brazilian ethanol production yeasts for maltose fermentation in media containing structurally complex nitrogen sources

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Sucrose Fermentation by Brazilian Ethanol Production Yeasts in Media Containing Structurally Complex Nitrogen Sources

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Influence of carbon and nitrogen sources on lipase production by a newly isolated Candida viswanathii strain

Microorganisms can produce lipases with different biochemical characteristics making necessary the screening of new lipase-producing strains for different industrial applications. In this study, 90 microbial strains were screened as potential lipase producers using a sensitive agar plate method with a suitable medium supplemented with Tween 20 and also a liquid culture supplemented with olive oil. The highest cell growth and lipase production for Candida viswanathii were observed in triolein and oleic acid when used as the only pure carbon source. Renewable low-cost triacylglycerols supported the best cell growth, and olive oil was found to be the best inducer for lipase production (19.50 g/L and 58.50 U). The selected conditions for enzyme production were found with yeast extract as nitrogen source and 1.5 % (w/v) olive oil (85.70 U) that resulted in a good cell growth yield (YX/S = 1.234 g/g) and lipase productivity (1.204 U/h) after 72 h of shake-flask cultivation. C. viswanathii lipase presented high hydrolytic activity on esters bonds of triacylglycerols of long-chain, and this strain can be considered an important candidate for future applications in chemical industries. © 2012 Springer-Verlag Berlin Heidelberg and the University of Milan.

Regulation of Sulfate Assimilation by Nitrogen in Arabidopsis

Using Arabidopsis, we analyzed the effect of omission of a nitrogen source and of the addition of different nitrogen-containing compounds on the extractable activity and the enzyme and mRNA accumulation of adenosine 5′-phosphosulfate reductase (APR). During 72 h without a nitrogen source, the APR activity decreased to 70% and 50% of controls in leaves and roots, respectively, while cysteine (Cys) and glutathione contents were not affected. Northern and western analysis revealed that the decrease of APR activity was correlated with decreased mRNA and enzyme levels. The reduced APR activity in roots could be fully restored within 24 h by the addition of 4 mM each of NO3 −, NH4 +, or glutamine (Gln), or 1 mM O-acetylserine (OAS). 35SO4 2− feeding showed that after addition of NH4 +, Gln, or OAS to nitrogen-starved plants, incorporation of 35S into proteins significantly increased in roots; however, glutathione and Cys labeling was higher only with Gln and OAS or with OAS alone, respectively. OAS strongly increased mRNA levels of all three APR isoforms in roots and also those of sulfite reductase, Cys synthase, and serine acetyltransferase. Our data demonstrate that sulfate reduction is regulated by nitrogen nutrition at the transcriptional level and that OAS plays a major role in this regulation.

Nitrogen-regulated Ubiquitination of the Gap1 Permease of Saccharomyces cerevisiae

Addition of ammonium ions to yeast cells growing on proline as the sole nitrogen source induces rapid inactivation and degradation of the general amino acid permease Gap1 through a process requiring the Npi1/Rsp5 ubiquitin (Ub) ligase. In this study, we show that NH4+ induces endocytosis of Gap1, which is then delivered into the vacuole where it is degraded. This down-regulation is accompanied by increased conversion of Gap1 to ubiquitinated forms. Ubiquitination and subsequent degradation of Gap1 are impaired in the npi1 strain. In this mutant, the amount of Npi1/Rsp5 Ub ligase is reduced >10-fold compared with wild-type cells. The C-terminal tail of Gap1 contains sequences, including a di-leucine motif, which are required for NH4+-induced internalization and degradation of the permease. We show here that mutant Gap1 permeases affected in these sequences still bind Ub. Furthermore, we provide evidence that only a small fraction of Gap1 is modified by Ub after addition of NH4+ to mutants defective in endocytosis.

Effects of nitrogen sources on the nitrate assimilation in Haloferax mediterranei: growth kinetics and transcriptomic analysis

The haloarchaeon Haloferax mediterranei is able to grow in a defined culture media not only in the presence of inorganic nitrogen salt but also with amino acid as the sole nitrogen source. Assimilatory nitrate and nitrite reductases, respectively, catalyze the first and second reactions. The genes involved in this process are nasA, which encodes nitrate reductase and is found within the operon nasABC, and nasD, which encodes nitrite reductase. These genes are subjected to transcriptional regulation, being repressed in the presence of ammonium and induced with either nitrate or nitrite. This type of regulation has also been described when the amino acids are used as nitrogen source in the minimal media. Furthermore, it has been observed that the microorganism growth depends on nitrogen source, obtaining the lowest growth rate in the presence of nitrate and aspartate. In this paper, we present the results of a comparative study of microorganism growth and transcriptomic analysis of the operon nasABC and gene nasD in different nitrogen sources. The results are the first ever produced in relation to amino acids as nitrogen sources within the Halobacteriaceae family.

Transcriptional profiles of Haloferax mediterranei based on nitrogen availability

The haloarchaeon Haloferax mediterranei is able to grow in the presence of different inorganic and organic nitrogen sources by means of the assimilatory pathway under aerobic conditions. In order to identify genes of potential importance in nitrogen metabolism and its regulation in the halophilic microorganism, we have analysed its global gene expression in three culture media with different nitrogen sources: (a) cells were grown stationary and exponentially in ammonium, (b) cells were grown exponentially in nitrate, and (c) cells were shifted to nitrogen starvation conditions. The main differences in the transcriptional profiles have been identified between the cultures with ammonium as nitrogen source and the cultures with nitrate or nitrogen starvation, supporting previous results which indicate the absence of ammonium as the factor responsible for the expression of genes involved in nitrate assimilation pathway. The results have also permitted the identification of transcriptional regulators and changes in metabolic pathways related to the catabolism and anabolism of amino acids or nucleotides. The microarray data was validated by real-time quantitative PCR on 4 selected genes involved in nitrogen metabolism. This work represents the first transcriptional profiles study related to nitrogen assimilation metabolism in extreme halophilic microorganisms using microarray technology.

Cross-talk towards the response regulator NtrC controlling nitrogen metabolism in Rhodobacter capsulatus

Rhodobacter capsulatus NtrB/NtrC two-component regulatory system controls expression of genes involved in nitrogen metabolism including urease and nitrogen fixation genes. The ntrY-ntrX genes, which are located immediately downstream of the nifR3-ntrB-ntrC operon, code for a two-component system of unknown function. Transcription of ntrY starts within the ntrC-ntrY intergenic region as shown by primer extension analysis, but maximal transcription requires, in addition, the promoter of the nifR3-ntrB-ntrC operon. While ntrB and ntrY single mutant strains were able to grow with either urea or N-2 as sole nitrogen source, a ntrB/ntrY double mutant (like a ntrC-deficient strain) was no longer able to use urea or N-2. These findings suggest that the histidine kinases NtrB and NtrY can substitute for each other as phosphodonors towards the response regulator NtrC.

(Table 1) Stable nitrogen isotopes of ammonium in interstitial waters from ODP Site 164-997

Ammonium (NH4+) concentration profiles in piston-core sediments of the Carolina Rise and Blake Ridge generally have linear concentration profiles within the sulfate reduction zone (Borowski, 1998). Deep Sea Drilling Project (DSDP) Site 533, located on the Blake Ridge, also displayed a linear ammonium concentration profile through the sulfate reduction zone and the profile linearity continues into the upper methanogenic zone to a depth of ~200 meters below seafloor (mbsf), where the first methane gas hydrates probably occur (Jenden and Gieskes, 1983, doi:10.2973/dsdp.proc.76.114.1983; Kvenvolden and Barnard, 1983, doi:10.2973/dsdp.proc.76.106.1983). Sediments from the Ocean Drilling Program (ODP) Leg 164 deep holes (Sites 994, 995, and 997) also exhibit linear ammonium profiles above the top of the gas hydrate zone (~200 mbsf) (Paull, Matsumoto, Wallace, et al., 1996, doi:10.2973/odp.proc.ir.164.1996). We hypothesized that a possible cause of linear ammonium profiles was diffusion of ammonium from a concentrated ammonium source at depth. We further reasoned that if this ammonium were produced by microbial fermentation reactions at depth, that a comparison of the nitrogen isotopic composition of sedimentary organic nitrogen and the nitrogen with pore-water ammonium would test this hypothesis. Convergence with depth of d15N values of the nitrogen source (sedimentary organic matter) and the nitrogen product (dissolved NH4+) would strongly suggest that ammonium was produced within a particular depth zone by microbial fermentation reactions. Here, we report d15N values of pore-water ammonium from selected interstitial water (IW) samples from Site 997, sampled during ODP Leg 164.

Effect of nitrogen nutrition on yeast ecology and alcoholic fermentation

Mestrado Vinifera Euromaster - Instituto Superior de Agronomia - UL

Use of Bioscreen C for growth of Mucor hiemalis in different carbon and nitrogen sources

O sistema automatizado Bioscreen C foi utilizado para o crescimento de quatro linhagens de Mucor hiemalis, isoladas do solo da Estação Ecológica de Juréia-Itatins (EEJI), estado de São Paulo, em meios líquidos com uma única fonte de carbono (2%) ou de nitrogênio (1%), pH 5,0, a 25ºC, e agitação de 150rpm por 5 dias. O meio com somente uma única fonte de nitrogênio foi adicionado com 2% de glicose. As leituras de densidade óptica foram realizadas a 540nm, em intervalos de 2h, por cinco dias. Os resultados foram analisados estatisticamente com o Teste de Friedman (alfa = 5%). Os melhores crescimentos foram obtidos com as linhagens M1, M2 e M3, que atingiram o início da fase log em 60 horas de cultivo. As melhores fontes de carbono variaram de acordo com a linhagem estudada, e extrato de levedura provou ser a melhor fonte de nitrogênio para todas as linhagens. Acetato de sódio inibiu o crescimento das quatro linhagens, sendo a M3 a mais afetada. O uso do sistema automatizado foi muito conveniente para as culturas em meio liquido, sendo rápido e automático, constituindo em uma boa técnica para a determinação das condições ambientais ótimas para crescimento de fungos filamentosos.

Activated carbons with high nitrogen content by a combination of hydrothermal carbonization with activation

This paper reports the production of carbons materials with a nitrogen content around 8%(w/w) and a well-developed porous structure, with BET surface area and pore volume up to 2130 m2 g−1 and 1.12 cm3 g−1, respectively, produced by a combination of hydrothermal carbonization, an environmental friendly method in the production of sustainable tunable carbon materials, with traditional activation methods. The porosity was developed through an activation process according to different routes, namely activation with CO2 and chemical activation using CaCO3 and K2CO3. The successful production of activated carbons using chitosan as a nitrogen source revealed to be a good alternative to post-synthesis methods.

Xylitol production from DEO hydrolysate of corn stover by Pichia stipitis YS-30

Corn stover that had been treated with vapor-phase diethyl oxalate released a mixture of mono- and oligosaccharides consisting mainly of xylose and glucose. Following overliming and neutralization, a d-xylulokinase mutant of Pichia stipitis, FPL-YS30 (xyl3-a dagger 1), converted the stover hydrolysate into xylitol. This research examined the effects of phosphoric or gluconic acids used for neutralization and urea or ammonium sulfate used as nitrogen sources. Phosphoric acid improved color and removal of phenolic compounds. d-Gluconic acid enhanced cell growth. Ammonium sulfate increased cell yield and maximum specific cell growth rate independently of the acid used for neutralization. The highest xylitol yield (0.61 g(xylitol)/g(xylose)) and volumetric productivity (0.18 g(xylitol)/g(xylose) l) were obtained in hydrolysate neutralized with phosphoric acid. However, when urea was the nitrogen source the cell yield was less than half of that obtained with ammonium sulfate.

Influence of ammonium sulphate feeding time on fed-batch Arthrospira (Spirulina) platensis cultivation and biomass composition with and without pH control

Previous work demonstrated that a mixture of NH(4)Cl and KNO(3) as nitrogen source was beneficial to fed-batch Arthrospira (Spirulina) platensis cultivation, in terms of either lower costs or higher cell concentration. On the basis of those results, this study focused on the use of a cheaper nitrogen source mixture, namely (NH(4))(2)SO(4) plus NaNO(3), varying the ammonium feeding time (T = 7-15 days), either controlling the pH by CO(2) addition or not. A. platensis was cultivated in mini-tanks at 30 degrees C, 156 mu mol photons m(-2) s(-1), and starting cell concentration of 400 mg L(-1), on a modified Schlosser medium. T = 13 days under pH control were selected as optimum conditions, ensuring the best results in terms of biomass production (maximum cell concentration of 2911 mg L(-1), cell productivity of 179 mg L(-1) d(-1) and specific growth rate of 0.77 d(-1)) and satisfactory protein and lipid contents (around 30% each). (C) 2011 Elsevier Ltd. All rights reserved.