Re-oxidation of cytosolic NADH is a major contributor to the high oxygen requirements of the thermotolerant yeast Ogataea parapolymorpha in oxygen-limited cultures

Thermotolerance is an attractive feature for yeast-based industrial ethanol production. However, incompletely understood oxygen requirements of known thermotolerant yeasts are incompatible with process requirements. To study the magnitude and molecular basis of these oxygen requirements in the facultatively fermentative, thermotolerant yeast Ogataea parapolymorpha, chemostat studies were performed under defined oxygen-sufficient and oxygen-limited cultivation regimes. The minimum oxygen requirements of O. parapolymorpha were found to be at least an order of magnitude larger than those of the thermotolerant yeast Kluyveromyces marxianus. This high oxygen requirement coincided with absence of glycerol formation, which plays a key role in NADH reoxidation in oxygen-limited cultures of other facultatively fermentative yeasts. Co-feeding of acetoin, whose reduction to 2,3-butanediol can reoxidize cytosolic NADH, supported a 2.5-fold higher biomass concentration in oxygen-limited cultures. The apparent inability of O. parapolymorpha to produce glycerol correlated with absence of orthologs of the S. cerevisiae genes encoding glycerol-3P phosphatase (ScGPP1, ScGPP2). Glycerol production was observed in aerobic batch cultures of a strain in which genes including key enzymes in mitochondrial reoxidation of NADH were deleted. However, transcriptome analysis did not identify a clear candidate for the responsible phosphatase. Expression of ScGPD2, encoding NAD+-dependent glycerol-3P dehydrogenase, and ScGPP1 in O. parapolymorpha resulted in increased glycerol production in oxygen-limited chemostats, but glycerol production rates remained substantially lower than observed in S. cerevisiae and K. marxianus. These results identify a dependency on aerobic respiration for reoxidation of NADH generated in biosynthesis as a key factor in the unexpectedly high oxygen requirements of O. parapolymorpha.

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Competition between Metals for Binding to Methanobactin Enables Expression of Soluble Methane Monooxygenase in the Presence of Copper

Applied and Environmental Microbiology ◽

10.1128/aem.03151-14 ◽

2014 ◽

Vol 81 (3) ◽

pp. 1024-1031 ◽

Cited By ~ 16

Author(s):

Bhagyalakshmi Kalidass ◽

Muhammad Farhan Ul-Haque ◽

Bipin S. Baral ◽

Alan A. DiSpirito ◽

Jeremy D. Semrau

Keyword(s):

Methane Monooxygenase ◽

High Specificity ◽

Copper Uptake ◽

Content Type ◽

Soluble Methane Monooxygenase ◽

Sds Page ◽

Genes Encoding ◽

Key Factor ◽

Membrane Bound

ABSTRACTIt is well known that copper is a key factor regulating expression of the two forms of methane monooxygenase found in proteobacterial methanotrophs. Of these forms, the cytoplasmic, or soluble, methane monooxygenase (sMMO) is expressed only at low copper concentrations. The membrane-bound, or particulate, methane monooxygenase (pMMO) is constitutively expressed with respect to copper, and such expression increases with increasing copper. Recent findings have shown that copper uptake is mediated by a modified polypeptide, or chalkophore, termed methanobactin. Although methanobactin has high specificity for copper, it can bind other metals, e.g., gold. Here we show that inMethylosinus trichosporiumOB3b, sMMO is expressed and active in the presence of copper if gold is also simultaneously present. Such expression appears to be due to gold binding to methanobactin produced byM. trichosporiumOB3b, thereby limiting copper uptake. Such expression and activity, however, was significantly reduced if methanobactin preloaded with copper was also added. Further, quantitative reverse transcriptase PCR (RT-qPCR) of transcripts of genes encoding polypeptides of both forms of MMO and SDS-PAGE results indicate that both sMMO and pMMO can be expressed when copper and gold are present, as gold effectively competes with copper for binding to methanobactin. Such findings suggest that under certain geochemical conditions, both forms of MMO may be expressed and activein situ. Finally, these findings also suggest strategies whereby field sites can be manipulated to enhance sMMO expression, i.e., through the addition of a metal that can compete with copper for binding to methanobactin.

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Regulation of Major Cellulosomal Endoglucanases of Clostridium thermocellum Differs from That of a Prominent Cellulosomal Xylanase

Journal of Bacteriology ◽

10.1128/jb.187.7.2261-2266.2005 ◽

2005 ◽

Vol 187 (7) ◽

pp. 2261-2266 ◽

Cited By ~ 37

Author(s):

Tali W. Dror ◽

Adi Rolider ◽

Edward A. Bayer ◽

Raphael Lamed ◽

Yuval Shoham

Keyword(s):

Growth Rate ◽

Clostridium Thermocellum ◽

Transcript Level ◽

Clear Correlation ◽

Transcript Levels ◽

Batch Cultures ◽

Processive Endoglucanase ◽

The Family ◽

Genes Encoding ◽

Family 10 Xylanase

ABSTRACT The expression of scaffoldin-anchoring genes and one of the major processive endoglucanases (CelS) from the cellulosome of Clostridium thermocellum has been shown to be dependent on the growth rate. For the present work, we studied the gene regulation of selected cellulosomal endoglucanases and a major xylanase in order to examine the previously observed substrate-linked alterations in cellulosome composition. For this purpose, the transcript levels of genes encoding endoglucanases CelB, CelG, and CelD and the family 10 xylanase XynC were determined in batch cultures, grown on either cellobiose or cellulose, and in carbon-limited continuous cultures at different dilution rates. Under all conditions tested, the transcript levels of celB and celG were at least 10-fold higher than that of celD. Like the major processive endoglucanase CelS, the transcript levels of these endoglucanase genes were also dependent on the growth rate. Thus, at a rate of 0.04 h−1, the levels of celB, celG, and celD were threefold higher than those obtained in cultures grown at maximal rates (0.35 h−1) on cellobiose. In contrast, no clear correlation was observed between the transcript level of xynC and the growth rate—the levels remained relatively high, fluctuating between 30 and 50 transcripts per cell. The results suggest that the regulation of C. thermocellum endoglucanases is similar to that of the processive endoglucanase celS but differs from that of a major cellulosomal xylanase in that expression of the latter enzyme is independent of the growth rate.

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Truncated Prosequence of Rhizopus oryzae Lipase: Key Factor for Production Improvement and Biocatalyst Stability

Catalysts ◽

10.3390/catal9110961 ◽

2019 ◽

Vol 9 (11) ◽

pp. 961 ◽

Cited By ~ 2

Author(s):

Josu López-Fernández ◽

Juan J. Barrero ◽

M. Dolors Benaiges ◽

Francisco Valero

Keyword(s):

Amino Acids ◽

Ionic Strength ◽

Rhizopus Oryzae ◽

Lipase Production ◽

Biochemical Properties ◽

Mature Sequence ◽

Rhizopus Oryzae Lipase ◽

Batch Cultures ◽

Production Improvement ◽

Key Factor

Recombinant Rhizopus oryzae lipase (mature sequence, rROL) was modified by adding to its N-terminal 28 additional amino acids from the C-terminal of the prosequence (proROL) to obtain a biocatalyst more suitable for the biodiesel industry. Both enzymes were expressed in Pichia pastoris and compared in terms of production bioprocess parameters, biochemical properties, and stability. Growth kinetics, production, and yields were better for proROL harboring strain than rROL one in batch cultures. When different fed-batch strategies were applied, lipase production and volumetric productivity of proROL-strain were always higher (5.4 and 4.4-fold, respectively) in the best case. rROL and proROL enzymatic activity was dependent on ionic strength and peaked in 200 mM Tris-HCl buffer. The optimum temperature and pH for rROL were influenced by ionic strength, but those for proROL were not. The presence of these amino acids altered lipase substrate specificity and increased proROL stability when different temperature, pH, and methanol/ethanol concentrations were employed. The 28 amino acids were found to be preferably removed by proteases, leading to the transformation of proROL into rROL. Nevertheless, the truncated prosequence enhanced Rhizopus oryzae lipase heterologous production and stability, making it more appropriate as industrial biocatalyst.

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Characterization and Expression of Genes Encoding Superoxide Dismutase in the Oriental Armyworm, Mythimna separata (Lepidoptera: Noctuidae)

Journal of Economic Entomology ◽

10.1093/jee/toz163 ◽

2019 ◽

Vol 112 (5) ◽

pp. 2381-2388 ◽

Cited By ~ 1

Author(s):

Hong-Bo Li ◽

Chang-Geng Dai ◽

Yong-Fu He ◽

Yang Hu

Keyword(s):

Superoxide Dismutase ◽

Population Density ◽

Developmental Stages ◽

Mythimna Separata ◽

Expression Of Genes ◽

Genes Encoding ◽

Key Factor ◽

Important Pest ◽

Oriental Armyworm

Abstract Superoxide dismutase (SOD) is an antioxidant metalloenzyme that catalyzes the dismutation of the superoxide anion O2− to O2 and H2O2. Many studies have focused on the role of SOD in response to abiotic stress, but its role during biotic stress, such as changes in organismal population density, has rarely been investigated. The oriental armyworm, Mythimna separata, is an economically important pest that exhibits phenotypic changes in response to population density. Solitary and gregarious phases occur at low and high population density, respectively. To examine the role of SODs in response to population density stress, we cloned two genes encoding SOD, MsCuZnSOD and MsMnSOD, and compared their expression in solitary and gregarious phases of M. separata. The MsCuZnSOD and MsMnSOD ORFs were 480 and 651 bp and encoded predicted protein products of 159 and 216 amino acids, respectively. The two SODs contained motifs that are typical of orthologous proteins. Real-time PCR indicated that the two SOD genes were expressed throughout developmental stages and were significantly upregulated in more mature stages of gregarious M. separata. Expression of the two SOD genes in various tissues of sixth-instar larvae was higher in gregarious versus solitary insects. Furthermore, expression of the SOD genes was significantly upregulated in response to crowding in solitary individuals, but suppressed in gregarious insects subjected to isolation. Collectively, these results suggest that population density may be key factor in the induction of SOD genes in M. separata.

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Second generation bioethanol production at high gravity of pilot-scale pretreated wheat straw employing newly isolated thermotolerant yeast Kluyveromyces marxianus DBTIOC-35

RSC Advances ◽

10.1039/c5ra05792b ◽

2015 ◽

Vol 5 (47) ◽

pp. 37485-37494 ◽

Cited By ~ 18

Author(s):

Jitendra Kumar Saini ◽

Ruchi Agrawal ◽

Alok Satlewal ◽

Reetu Saini ◽

Ravi Gupta ◽

...

Keyword(s):

Kluyveromyces Marxianus ◽

Biomass Concentration ◽

Pilot Scale ◽

Process Time ◽

High Gravity ◽

Thermotolerant Yeast ◽

Second Generation Bioethanol ◽

P Application ◽

Pretreated Wheat Straw ◽

High Biomass Concentration

Application of thermotolerant yeast Kluyveromyces marxianus DBTIOC-35 in SSF decreases overall process time, and increases productivity and yield by allowing elimination of presaccharification step and use of high biomass concentration, respectively.

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The Effect of Hot Deformation on Dispersoid Evolution in a Model 3xxx Alloy

Materials Science Forum ◽

10.4028/www.scientific.net/msf.794-796.697 ◽

2014 ◽

Vol 794-796 ◽

pp. 697-703 ◽

Cited By ~ 4

Author(s):

Joseph D. Robson ◽

Thomas Hill ◽

Nicolas Kamp

Keyword(s):

Hot Deformation ◽

High Strain ◽

Volume Fraction ◽

Strain Rates ◽

Number Density ◽

Key Factor ◽

Order Of Magnitude ◽

Hot Torsion ◽

3Xxx Alloy ◽

Magnitude Increase

The influence of hot deformation on the evolution of size, shape, and fraction of dispersoids has been studied in a simple 3xxx aluminium alloy by means of hot torsion testing. It has been shown that at high strain rates, deformation leads to spheroidization of the dispersoids, an increase in number density, and an increase in volume fraction. The increase in number density and volume fraction are associated with precipitation of new particles. The enhancement of manganese diffusion is a key factor in promoting rapid dispersoid evolution during deformation. A model has been developed to estimate the effect of deformation induced vacancies and dislocations on diffusion. This predicts that an order of magnitude increase in diffusion coefficient between may occur under typical hot deformation conditions, consistent with the rapid microstructural changes measured experimentally.

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A histamine gene expression program in human macrophages.

10.31219/osf.io/stfup ◽

2020 ◽

Author(s):

Shahan Mamoor

Keyword(s):

Gene Expression ◽

Septic Shock ◽

Cell Types ◽

Gene Expression Program ◽

Dna And Rna ◽

Genes Encoding ◽

Unbiased Manner ◽

Order Of Magnitude ◽

Epigenetic Modulation ◽

Expression Program

The significance of histamine signaling in septic shock is debated (1-5). The macrophage is one of the critical cell types that provide rapid immune responses to early events in septic shock (6-11). To understand in a systematic and unbiased manner the transcriptional behavior of macrophage in response to the immunologic effector histamine, we used a public dataset (12) to describe the most significant changes in the gene expression program of human primary macrophages from response to histamine exposure. We found that of the genes whose expression was most different between macrophages exposed to histamine and the naïve macrophage there were a selection of the genes whose expression could be assigned to 22 discrete modules, including genes encoding enzymes involved in glycosylation, the mitochondria and metabolism, the quality control and translation of proteins, the ER stress response, the Rab family of molecules, genes that bind or encode molecules involved in genetic and epigenetic modulation of DNA and RNA, CXCL5, and the cyclin dependent kinase CDK14. It appears that macrophages respond to histamine exposure by altering gene expression at a relatively large number of loci but by a relatively small order of magnitude.

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