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1.
Background
The conversion of plant biomass to ethanol via enzymatic cellulose hydrolysis offers a potentially sustainable route to biofuel production. However, the inhibition of enzymatic activity in pretreated biomass by lignin severely limits the efficiency of this process.Results
By performing atomic-detail molecular dynamics simulation of a biomass model containing cellulose, lignin, and cellulases (TrCel7A), we elucidate detailed lignin inhibition mechanisms. We find that lignin binds preferentially both to the elements of cellulose to which the cellulases also preferentially bind (the hydrophobic faces) and also to the specific residues on the cellulose-binding module of the cellulase that are critical for cellulose binding of TrCel7A (Y466, Y492, and Y493).Conclusions
Lignin thus binds exactly where for industrial purposes it is least desired, providing a simple explanation of why hydrolysis yields increase with lignin removal.2.
Objectives
The biochemical conversion of lignocellulosic biomass into renewable fuels and chemicals provides new challenges for industrial scale processes. One such process, which has received little attention, but is of great importance for efficient product recovery, is solid–liquid separations, which may occur both after pretreatment and after the enzymatic hydrolysis steps. Due to the changing nature of the solid biomass during processing, the solid–liquid separation properties of the biomass can also change. The objective of this study was to show the effect of enzymatic hydrolysis of cellulose upon the water retention properties of pretreated biomass over the course of the hydrolysis reaction.Results
Water retention value measurements, coupled with 1H NMR T2 relaxometry data, showed an increase in water retention and constraint of water by the biomass with increasing levels of cellulose hydrolysis. This correlated with an increase in the fines fraction and a decrease in particle size, suggesting that structural decomposition rather than changes in chemical composition was the most dominant characteristic.Conclusions
With increased water retained by the insoluble fraction as cellulose hydrolysis proceeds, it may prove more difficult to efficiently separate hydrolysis residues from the liquid fraction with improved hydrolysis.3.
N. Cesbron A.-L. Royer Y. Guitton A. Sydor B. Le Bizec G. Dervilly-Pinel 《Metabolomics : Official journal of the Metabolomic Society》2017,13(8):99
Introduction
Collecting feces is easy. It offers direct outcome to endogenous and microbial metabolites.Objectives
In a context of lack of consensus about fecal sample preparation, especially in animal species, we developed a robust protocol allowing untargeted LC-HRMS fingerprinting.Methods
The conditions of extraction (quantity, preparation, solvents, dilutions) were investigated in bovine feces.Results
A rapid and simple protocol involving feces extraction with methanol (1/3, M/V) followed by centrifugation and a step filtration (10 kDa) was developed.Conclusion
The workflow generated repeatable and informative fingerprints for robust metabolome characterization.4.
Tânia M. Shiga Weihua Xiao Haibing Yang Ximing Zhang Anna T. Olek Bryon S. Donohoe Jiliang Liu Lee Makowski Tao Hou Maureen C. McCann Nicholas C. Carpita Nathan S. Mosier 《Biotechnology for biofuels》2017,10(1):310
Background
The crystallinity of cellulose is a principal factor limiting the efficient hydrolysis of biomass to fermentable sugars or direct catalytic conversion to biofuel components. We evaluated the impact of TFA-induced gelatinization of crystalline cellulose on enhancement of enzymatic digestion and catalytic conversion to biofuel substrates.Results
Low-temperature swelling of cotton linter cellulose in TFA at subzero temperatures followed by gentle heating to 55 °C dissolves the microfibril structure and forms composites of crystalline and amorphous gels upon addition of ethanol. The extent of gelatinization of crystalline cellulose was determined by reduction of birefringence in darkfield microscopy, loss of X-ray diffractability, and loss of resistance to acid hydrolysis. Upon freeze-drying, an additional degree of crystallinity returned as mostly cellulose II. Both enzymatic digestion with a commercial cellulase cocktail and maleic acid/AlCl3-catalyzed conversion to 5-hydroxymethylfurfural and levulinic acid were markedly enhanced with the low-temperature swollen cellulose. Only small improvements in rates and extent of hydrolysis and catalytic conversion were achieved upon heating to fully dissolve cellulose.Conclusions
Low-temperature swelling of cellulose in TFA substantially reduces recalcitrance of crystalline cellulose to both enzymatic digestion and catalytic conversion. In a closed system to prevent loss of fluorohydrocarbons, the relative ease of recovery and regeneration of TFA by distillation makes it a potentially useful agent in large-scale deconstruction of biomass, not only for enzymatic depolymerization but also for enhancing rates of catalytic conversion to biofuel components and useful bio-products.5.
Galina R. Demina Vadim D. Nikitushkin Margarita O. Shleeva Olga B. Riabova Alexander Yu. Lepioshkin Vadim A. Makarov Arseny S. Kaprelyants 《Annals of clinical microbiology and antimicrobials》2017,16(1):69
Background
Resuscitation promoting factors (Rpfs) are the proteins involved in the process of reactivation of the dormant cells of mycobacteria. Recently a new class of nitrophenylthiocyanates (NPTs), capable of inhibiting the biological and enzymatic activities of Rpfs has been discovered. In the current study the inhibitory properties of the compounds containing both nitro and thiocyanate groups alongside with the compounds with the modified number and different spatial location of the substituents are compared.Methods
New benzoylphenyl thiocyanates alongside with nitrophenylthiocyanates were tested in the enzymatic assay of bacterial peptidoglycan hydrolysis as well as against strains of several actinobacteria (Mycobacterium smegmatis, Mycobacterium tuberculosis) on in-lab developed models of resuscitation of the dormant forms.Results
Introduction of the additional nitro and thiocyanate groups to the benzophenone scaffold did not influence the inhibitory activity of the compounds. Removal of the nitro groups analogously did not impair the functional properties of the molecules. Among the tested compounds two molecules without nitro group: 3-benzoylphenyl thiocyanate and 4-benzoylphenyl thiocyanate demonstrated the maximum activity in both enzymatic assay (inhibition of the Rpf-mediated peptidoglycan hydrolysis) and in the resuscitation assay of the dormant M. tuberculosis cells.Conclusions
The current study demonstrates dispensability of the nitro group in the NPT’s structure for inhibition of the enzymatic and biological activities of the Rpf protein molecules. These findings provide new prospects in anti-TB drug discovery especially in finding of molecular scaffolds effective for the latent infection treatment.6.
Transient expression of recombinant tissue plasminogen activator (rt-PA) gene in cucurbit plants using viral vector 总被引:1,自引:0,他引:1
Vahid Jalali Javaran Alireza Shafeinia Mokhtar Jalali Javaran Esmaeil Ghasemi Gojani Malihe Mirzaee 《Biotechnology letters》2017,39(4):607-612
Objective
To use a transient expression system to express a truncated human tissue plasminogen activator (K2S) gene in cucurbit plants.Results
The recombinant tissue plasminogen activator protein (K2S form) was expressed in active form in cucurbit plants. Its molecular weight was 43 kDa. The plant-derived rt-PA was determined using goat anti-rabbit antibody by western blotting. Among the infected lines, the highest expression of rt-PA was 62 ng/100 mg per leaf tissue as measured by ELISA. The enzymatic activity of the plant-derived rt-PA was 0.8 IU/ml.Conclusions
The K25 form of rt-PA was expressed for the first time using the viral expression system. Plant-derived rt-PA showed similar potency to commercially-available PA.7.
Morten Ambye-Jensen Riccardo Balzarotti Sune Tjalfe Thomsen César Fonseca Zsófia Kádár 《Biotechnology for biofuels》2018,11(1):336
Background
Ensiling cannot be utilized as a stand-alone pretreatment for sugar-based biorefinery processes but, in combination with hydrothermal processing, it can enhance pretreatment while ensuring a stable long-term storage option for abundant but moist biomass. The effectiveness of combining ensiling with hydrothermal pretreatment depends on biomass nature, pretreatment, and silage conditions.Results
In the present study, the efficiency of the combined pretreatment was assessed by enzymatic hydrolysis and ethanol fermentation, and it was demonstrated that ensiling of sugarcane bagasse produces organic acids that can partly degrade biomass structure when in combination with hydrothermal treatment, with the consequent improvement of the enzymatic hydrolysis of cellulose and of the overall 2G bioethanol process efficiency. The optimal pretreatment conditions found in this study were those using ensiling and/or hydrothermal pretreatment at 190 °C for 10 min as this yielded the highest overall glucose recovery yield and ethanol yield from the raw material (0.28–0.30 g/g and 0.14 g/g, respectively).Conclusion
Ensiling prior to hydrothermal pretreatment offers a controlled solution for wet storage and long-term preservation for sugarcane bagasse, thus avoiding the need for drying. This preservation method combined with long-term storage practice can be an attractive option for integrated 1G/2G bioethanol plants, as it does not require large capital investments or energy inputs and leads to comparable or higher overall sugar recovery and ethanol yields.8.
Objectives
To use permeabilized cells of the fission yeast, Schizosaccharomyces pombe, that expresses human UDP-glucose 6-dehydrogenase (UGDH, EC 1.1.1.22), for the production of UDP-glucuronic acid from UDP-glucose.Results
In cell extracts no activity was detected. Therefore, cells were permeabilized with 0.3 % (v/v) Triton X-100. After washing away all low molecular weight metabolites, the permeabilized cells were directly used as whole cell biocatalyst. Substrates were 5 mM UDP-glucose and 10 mM NAD+. Divalent cations were not added to the reaction medium as they promoted UDP-glucose hydrolysis. With this reaction system 5 mM UDP-glucose were converted into 5 mM UDP-glucuronic acid within 3 h.Conclusions
Recombinant permeabilized cells of S. pombe can be used to synthesize UDP-glucuronic acid with 100 % yield and selectivity.9.
Matthew J. Roberts Clement W. K. Chow Martin Lavin Gregory K. Pierens Robert A. Gardiner 《Metabolomics : Official journal of the Metabolomic Society》2016,12(11):162
Introduction
Human seminal fluid (hSF) has been suggested as a biofluid suitable to characterise male reproductive organ pathology with metabolomics. However, various enzymatic processes, including phosphorylcholine hydrolysis mediated by prostatic acid phosphatase (PAP), cause unwanted metabolite variation that may complicate metabolomic analysis of fresh hSF samples.Objectives
To investigate the effects of PAP inhibition with tartrate.Methods
Using NMR spectroscopy, the kinetics of phosphorylcholine to choline hydrolysis was characterized in hSF samples from three subjects at different temperatures and tartrate concentrations. Principal components analysis was used to characterise the effects of tartrate and temperature on personal differences in metabolite profiles. Potential effects of tartrate on RNA quantification were also determined.Results
Metabolite profiles and the kinetics of phosphorylcholine degradation are reproducible in independent samples from three ostensibly normal subjects. Increasing concentrations of tartrate and refrigerated sample storage (279 K) resulted in greatly reduced reaction rates as judged by apparent rate constants. Multivariate statistical analysis showed that personal differences in metabolite profiles are not overshadowed by tartrate addition, which stabilises phosphorylcholine and choline concentrations. The tartrate signal also served as an internal concentration standard in the samples, allowing the determination of absolute metabolite concentrations in hSF. Furthermore, the presence of tartrate did not affect RNA expression analysis by qPCR.Conclusion
Based on these results we recommend as standard protocol for the collection of hSF samples, that 10 mM tartrate are added immediately to samples, followed by sample storage/handling at 277 K until clinical processing within 6 h to remove/inactivate enzymes and isolate metabolite supernatant and other cellular fractions.10.
Objectives
To optimize conversion of rutin to isoquercetin by commercial α-l-rhamnosidase using high hydrostatic pressure (HHP).Results
The de-rhamnosylation activity of α-l-rhamnosidase for isoquercetin production was maximal at pH 6.0 and 50 °C using HHP (150 MPa). The enzyme showed high specificity for rutin. The specific activity for rutin at HHP was 1.5-fold higher than that at atmospheric pressure. The enzyme completely hydrolysed 20 mM rutin in tartary buckwheat extract after 2 h at HHP, with a productivity of 10 mM h?1. The productivity and conversion were 2.2- and 1.5-fold higher at HHP than at atmospheric pressure, respectively.Conclusions
This is the first report concerning the enzymatic hydrolysis of isoquercetin in tartary buckwheat at HHP.11.
Background
Proteins play fundamental and crucial roles in nearly all biological processes, such as, enzymatic catalysis, signaling transduction, DNA and RNA synthesis, and embryonic development. It has been a long-standing goal in molecular biology to predict the tertiary structure of a protein from its primary amino acid sequence. From visual comparison, it was found that a 2D triangular lattice model can give a better structure modeling and prediction for proteins with short primary amino acid sequences.Methods
This paper proposes a hybrid of hill-climbing and genetic algorithm (HHGA) based on elite-based reproduction strategy for protein structure prediction on the 2D triangular lattice.Results
The simulation results show that the proposed HHGA can successfully deal with the protein structure prediction problems. Specifically, HHGA significantly outperforms conventional genetic algorithms and is comparable to the state-of-the-art method in terms of free energy.Conclusions
Thanks to the enhancement of local search on the global search, the proposed HHGA achieves promising results on the 2D triangular protein structure prediction problem. The satisfactory simulation results demonstrate the effectiveness of the proposed HHGA and the utility of the 2D triangular lattice model for protein structure prediction.12.
Rachel A. Spicer Christoph Steinbeck 《Metabolomics : Official journal of the Metabolomic Society》2018,14(1):16
Introduction
Data sharing is being increasingly required by journals and has been heralded as a solution to the ‘replication crisis’.Objectives
(i) Review data sharing policies of journals publishing the most metabolomics papers associated with open data and (ii) compare these journals’ policies to those that publish the most metabolomics papers.Methods
A PubMed search was used to identify metabolomics papers. Metabolomics data repositories were manually searched for linked publications.Results
Journals that support data sharing are not necessarily those with the most papers associated to open metabolomics data.Conclusion
Further efforts are required to improve data sharing in metabolomics.13.
Dimitrios J. Floros Paul R. Jensen Pieter C. Dorrestein Nobuhiro Koyama 《Metabolomics : Official journal of the Metabolomic Society》2016,12(9):145
Introduction
Natural products from culture collections have enormous impact in advancing discovery programs for metabolites of biotechnological importance. These discovery efforts rely on the metabolomic characterization of strain collections.Objective
Many emerging approaches compare metabolomic profiles of such collections, but few enable the analysis and prioritization of thousands of samples from diverse organisms while delivering chemistry specific read outs.Method
In this work we utilize untargeted LC–MS/MS based metabolomics together with molecular networking to inventory the chemistries associated with 1000 marine microorganisms.Result
This approach annotated 76 molecular families (a spectral match rate of 28 %), including clinically and biotechnologically important molecules such as valinomycin, actinomycin D, and desferrioxamine E. Targeting a molecular family produced primarily by one microorganism led to the isolation and structure elucidation of two new molecules designated maridric acids A and B.Conclusion
Molecular networking guided exploration of large culture collections allows for rapid dereplication of know molecules and can highlight producers of uniques metabolites. These methods, together with large culture collections and growing databases, allow for data driven strain prioritization with a focus on novel chemistries.14.
Objective
To examine the activities of residual enzymes in dried shiitake mushrooms, which are a traditional foodstuff in Japanese cuisine, for possible applications in food processing.Results
Polysaccharide-degrading enzymes remained intact in dried shiitake mushrooms and the activities of amylase, β-glucosidase and pectinase were high. A potato digestion was tested using dried shiitake powder. The enzymes reacted with potato tuber specimens to solubilize sugars even under a heterogeneous solid-state condition and that their reaction modes were different at 38 and 50 °C.Conclusion
Dried shiitake mushrooms have a potential use in food processing as an enzyme preparation.15.
Anita H. Lewin Peter Silinski James Hayes Amanda Gilbert S. Wayne Mascarella Herbert H. Seltzman 《Metabolomics : Official journal of the Metabolomic Society》2017,13(10):117
Introduction
Metabolomics analysis depends on the identification and validation of specific metabolites. This task is significantly hampered by the absence of well-characterized reference standards. The one-carbon carrier 10-formyltetrahydrofolate acts as a donor of formyl groups in anabolism, where it is a substrate in formyltransferase reactions in purine biosynthesis. It has been reported as an unstable substance and is currently unavailable as a reference standard for metabolomics analysis.Objectives
The current study was undertaken to provide the metabolomics community thoroughly characterized 10-formyltetrahydrofolate along with analytical methodology and guidelines for its storage and handling.Methods
Anaerobic base treatment of 5,10-methenyltetrahydrofolate chloride in the presence of antioxidant was utilized to prepare 10-formyltetrahydrofolate.Results
Pure 10-formyltetrahydrofolate has been prepared and physicochemically characterized. Conditions toward maintaining the stability of a solution of the dipotassium salt of 10-formyltetrahydrofolate have been determined.Conclusion
This study describes the facile preparation of pure (>90%) 10-formyltetrahydrofolate, its qualitative physicochemical characterization, as well as conditions to enable its use as a reference standard in physiologic samples.16.
Soil microbial activity in relation to dissolved organic matter properties under different tree species 总被引:1,自引:0,他引:1
Background and aims
The total concentration of dissolved organic carbon (DOC) has often been observed to correlate positively with soil microbial respiration. The aim was to explain the correlation with the properties of dissolved organic matter (DOM).Methods
A dataset from previously published papers was gathered together and subjected to multivariate analyses. Samples were collected from five tree species experiments in Finland. The degradability of DOM was assessed by measuring bacterial and fungal growth in DOM. The chemical properties of DOM were assessed by XAD resin fractionation and molecular weight. Soil microbial activity was assessed as C and N mineralization and microbial biomass.Results
Both low and high molecular weight compounds, as well as hydrophilic neutral compounds, seemed to be relatively easily degradable. In contrast to our presupposition, easily degradable DOM seemed to be less abundant in soil where variables describing microbial activity were higher. Birch soil with higher microbial biomass N seemed to contain less easily degradable DOM than spruce soil.Conclusion
We suggest that DOM collected and characterized at a certain point reflects more the accumulation of refractory compounds following high microbial activity than the easily degradable compounds that microbes would be using when measured.17.
Andrei Prodan Sultan Imangaliyev Henk S. Brand Martijn N. A. Rosema Evgeni Levin Wim Crielaard Bart J. F. Keijser Enno C. I. Veerman 《Metabolomics : Official journal of the Metabolomic Society》2016,12(9):147
Introduction
Understanding the changes occurring in the oral ecosystem during development of gingivitis could help improve prevention and treatment strategies for oral health. Erythritol is a non-caloric polyol proposed to have beneficial effects on oral health.Objectives
To examine the effect of experimental gingivitis and the effect of erythritol on the salivary metabolome and salivary functional biochemistry.Methods
In a two-week experimental gingivitis challenge intervention study, non-targeted, mass spectrometry-based metabolomic profiling was performed on saliva samples from 61 healthy adults, collected at five time-points. The effect of erythritol was studied in a randomized, controlled trial setting. Fourteen salivary biochemistry variables were measured with antibody- or enzymatic activity-based assays.Results
Bacterial amino acid catabolites (cadaverine, N-acetylcadaverine, and α-hydroxyisovalerate) and end-products of bacterial alkali-producing pathways (N-α-acetylornithine and γ-aminobutyrate) increased significantly during the experimental gingivitis. Significant changes were found in a set of 13 salivary metabolite ratios composed of host cell membrane lipids involved in cell signaling, host responses to bacteria, and defense against free radicals. An increase in mevalonate was also observed. There were no significant effects of erythritol. No significant changes were found in functional salivary biochemistry.Conclusions
The findings underline a dynamic interaction between the host and the oral microbial biofilm during an experimental induction of gingivitis.18.
Background
Mixtures of beta distributions are a flexible tool for modeling data with values on the unit interval, such as methylation levels. However, maximum likelihood parameter estimation with beta distributions suffers from problems because of singularities in the log-likelihood function if some observations take the values 0 or 1.Methods
While ad-hoc corrections have been proposed to mitigate this problem, we propose a different approach to parameter estimation for beta mixtures where such problems do not arise in the first place. Our algorithm combines latent variables with the method of moments instead of maximum likelihood, which has computational advantages over the popular EM algorithm.Results
As an application, we demonstrate that methylation state classification is more accurate when using adaptive thresholds from beta mixtures than non-adaptive thresholds on observed methylation levels. We also demonstrate that we can accurately infer the number of mixture components.Conclusions
The hybrid algorithm between likelihood-based component un-mixing and moment-based parameter estimation is a robust and efficient method for beta mixture estimation. We provide an implementation of the method (“betamix”) as open source software under the MIT license.19.
César Nicolás Juan P. Almeida Magnus Ellström Adam Bahr Sharon E. Bone Nicholas P. Rosenstock John R. Bargar Anders Tunlid Per Persson Håkan Wallander 《Plant and Soil》2017,414(1-2):113-125
Background and aims
Mineralization of soil organic matter (SOM) constitutes a major carbon flux to the atmosphere. The carbon use efficiency (CUE) of the saprotrophic microorganisms mineralizing SOM is integral for soil carbon dynamics. Here we investigate how the CUE is affected by temperature, metabolic conditions, and the molecular complexity of the substrate.Methods
We incubated O-horizon soil samples (with either 13C–glucose or 13C–cellulose) from a boreal coniferous forest at 4, 9, 14, and 19 °C, and calculated CUEs based on the amount of 13C–CO2 and 13C–labelled microbial biomass produced. The effects of substrate, temperature, and metabolic conditions (representing unlimited substrate supply and substrate limitation) on CUE were evaluated.Results
CUE from metabolizing glucose was higher as compared to cellulose. A slight decrease in CUE with increasing temperature was observed in glucose amended samples (but only in the range 9–19 °C), but not in cellulose amended samples. CUE differed significantly with metabolic conditions, i.e. CUE was higher during unlimited growth conditions as compared to conditions with substrate limitation.Conclusions
We conclude that it is integral to account for both differences in CUE during different metabolic phases, as well as complexity of substrate, when interpreting temperature dependence on CUE in incubation studies.20.
Jie Yang Jianhua Cheng Bo Sun Haijing Li Shengming Wu Fangting Dong Xianzhong Yan 《Metabolomics : Official journal of the Metabolomic Society》2018,14(4):40