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Glucose isomerase from Streptomyces murinus has been crystallized in space group P4(1)2(1)2, cell dimensions a = b = 137.65 and c = 132.20 A. One dimer of the tetrametric molecule is found per asymmetric unit. An initial structure solution was obtained by the molecular replacement method. The crystallographic refinement was performed using molecular dynamics techniques with X-ray restraints. The final crystallographic R value is 21.4% at 2.6 A resolution including 3023 non-H atoms, two metal ions and two water molecules per monomer.

Copyright of Journal of Phytopathology is the property of Wiley-Blackwell and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

In the field of our research interest, we designed new sulfa drugs–substituted norbornyl imides as prospective bioactive scaffolds by the reaction of endo‐endic anhydride with sulfa drugs followed by reductive Heck reactions of these products. Norbornenyl imides, as starting compounds, and their Heck products were achieved in good yields. Molecular characterization and stereochemistry of the compounds were investigated using Fourier transform infrared, liquid chromatography time‐of‐flight mass spectrometry, proton nuclear magnetic resonance, carbon 13 attached proton test, H‐H correlation spevtroscopy, and nuclear overhauser enhancement spectroscopy experiments. The Heck products of norbornenyl imides showed exo‐selectivity. All eight novel synthesized compounds, including two sulfa drugs–based norbornenyl imides and six exo‐5‐aryl‐ substituted norbornyl imides, as well as standard sulfonamides, were evaluated for antimicrobial activity against nine microorganisms consisting of Gram‐negative bacteria, Gram‐positive nonactinobacteria, actinobacteria, and yeast. The most active compound was 3a against actinobacteria. The efficacy of 2 and its derivatives against the studied strains were weaker than 3 and its derivatives, but all the studied samples exhibited antifungal potency. [ABSTRACT FROM AUTHOR]

BACKGROUND: Aldo‐keto isomerizations of D‐xylose and D‐glucose catalyzed by xylose isomerase (XI) is commercially carried out to produce bio‐ethanol and high fructose corn syrup. XI is a metalloenzyme. In this paper, kinetics and thermodynamics of the activity and stability of hyperthermophilic recombinant XI from Thermotoga naphthophila RKU‐10T (TnapXI) were studied in the presence and absence of metal ions. RESULTS: Kinetics parameters of recombinant XI from TnapXI were calculated as Km = 0.99 mmol L–1, Vmax = 500 μmol.mg−1.min−1, and kcat = 68.3 min−1 for the isomerization of D‐xylose(aldo)⇌D‐xylulose(keto), whereas for D‐glucose(aldo)⇌D‐fructose(keto) conversion Km = 7.72 mmol L–1, Vmax = 90 μmol.mg−1.min−1, and kcat = 12.4 min−1. Ionization constants were calculated as pKa1 = 6.0 and pKa2 = 7.6, whereas activation constants (Ka) of TnapXI were 0.3, 0.5, and 4.0 mmol L–1 for Co2+, Mn2+, and Mg2+, respectively. Inhibition constant (Ki) was 0.119 mmol L–1 for Ca2+. Q10 was 2.8 and Ea = 82.25 kJ.mol−1. Thermodynamics for isomerization were calculated as ∆H* = 79.2 kJ.mol−1, ∆G* = 83 kJ.mol−1, and ∆S* = −10.5 J.mol−1.K−1. The z‐value was 12.6 °C for apo‐TnapXI and 32.6 °C for holo‐TnapXI. Deactivation parameters were ∆G*(d) = 100 kJ.mol−1, ∆H*(d) = 206 kJ.mol−1, ∆S*(d) = 0.28 kJ.mol−1.K−1 for apo‐TnapXI and t1/2 = 18 min at 95 °C, while ∆G*(d) = 104 kJ.mol−1, ∆H*(d) = 767 kJ.mol−1, ∆S*(d) = 1.8 kJ.mol−1.K−1 for holo‐TnapXI and t1/2 = 65 min at 95 °C. CONCLUSION: These noteworthy features make this enzyme an attractive candidate for the industrial production of fuel ethanol and high fructose corn syrup at high temperatures to maximize the product yield. © 2021 Society of Chemical Industry (SCI). [ABSTRACT FROM AUTHOR]

Glucose isomerase (GI) that catalyzes the conversion of D-glucose to D-fructose is one of the most important industrial enzymes for the production of high-fructose corn syrup (HFCS). In this study, a novel GI (CbGI) was cloned from Caldicellulosiruptor bescii and expressed in Escherichia coli. The purified recombinant CbGI (rCbGI) showed neutral and thermophilic properties. It had optimal activities at pH 7.0 and 80°C and retained stability at 85°C. In comparison with other reported GIs, rCbGI exhibited higher substrate affinity (K m = 42.61 mM) and greater conversion efficiency (up to 57.3% with 3M D-glucose as the substrate). The high catalytic efficiency and affinity of this CbGI is much valuable for the cost-effective production of HFCS. [ABSTRACT FROM AUTHOR]

Recently immobilized enzymes have been widely used in industrial processes due to their outstanding advantages, such as high stability and recyclability; however, their kinetic behaviour is generally controlled by mass diffusion effects. Thus, in order to improve these enzymatic processes, a clear discernment between the kinetic and diffusion mechanisms that control the production of the metabolite require investigation. In practice, it is typical to establish apparent kinetics for immobilized enzyme operations, and the validity of the apparent kinetics is restricted to the studied cases. In this work, a new approach for mathematically describing the kinetic and diffusion mechanics in an immobilized biocatalyst bead is established, in which the fraction of residual enzymatic activity is included, and is defined as a measure of the active and available enzymes in the bead porous network. In addition, the diffusion and kinetic mechanisms are described by the effective diffusion coefficient and the free enzyme kinetics, since the porous network of the bead is assumed as the bioreaction volume. Therefore, free enzyme kinetics were determined from glucose to fructose bioconversion using a stirred tank reactor with free glucose‐isomerase, in which substrate and enzyme concentrations and temperature were varied. The fraction of residual enzymatic activity (η=0.553) and the effective diffusion coefficient (Deff=8.356×10−12m2/s) were obtained from the isomerization of glucose to fructose using a stirred tank reactor with immobilized glucose‐isomerase in calcium alginate beads at different substrate and enzyme concentrations. Finally, simulations were carried out to establish the bioreaction solid‐phase characteristics that most significantly influence productivity. [ABSTRACT FROM AUTHOR]

Copyright of EPPO Bulletin is the property of Wiley-Blackwell and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Green hydrogen production through water splitting at low temperatures is highly desired for hydrogen economy. Herein, we demonstrate an in vitro non-natural enzymatic pathway to utilize the chemical energy stored in xylooligosaccharides from biomass to split water to produce a nearly theoretical yield of H2 (i.e., ≈9.5 H2 per xylose plus water). This pathway was constructed on the basis of the novel activities of phosphopentomutase catalyzing the conversion of d-xylose 1-phosphate into d-xylose 5-phosphate and of ribose 5-phosphate isomerase catalyzing the conversions of d-xylose 5-phosphate and d-xylulose 5-phosphate. This study suggests that the discovery of novel promiscuous enzyme activities is important to implement complicated biotransformations catalyzed by synthetic enzymatic pathways. [ABSTRACT FROM AUTHOR]

Production of a xylose isomerase (XI) with high tolerance to the inhibitors xylitol and calcium, and high activity at the low pH and temperature conditions characteristic of yeast fermentations, is desirable for a simultaneous isomerization/fermentation process for cellulosic ethanol production. A putative XI gene (xylA) from the marine bacterium Fulvimarina pelagi was identified by sequence analysis of the F. pelagi genome, and was PCR amplified, cloned, and expressed in Escherichia coli. The rXI was produced in shake flask and fed-batch fermentations using glucose as the growth substrate. The optimum pH for rXI was approximately 7, although activity was evident at pH as low as 5.5. The purified rXI had a molecular weight in 160 kDA, a Vmax of 0.142 U/mg purified rXI, and a KM for xylose in the range of 1.75-4.17 mM/L at pH 6.5 and a temperature of 35°C. The estimated calcium and xylitol KI values for rXI in cell-free extracts were 2,500 mg/L and >50 mM, respectively. The low KM of the F. pelagi xylose isomerase is consistent with the low nutrient conditions of the pelagic environment. These results indicate that Ca2+ and xylitol are not likely to be inhibitory in applications employing the rXI from F. pelagi to convert xylose to xylulose in fermentations of complex biomass hydrolysates. A higher Vmax at low pH (<6) and temperature (30°C) would be preferable for use in biofuels production. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1230-1237, 2016 [ABSTRACT FROM AUTHOR]

Sugar phosphates cannot be produced easily by microbial fermentation because negatively-charged compounds cannot be secreted across intact cell membrane. D-xylulose 5-phosphate (Xu5P), a very expensive sugar phosphate, was synthesized from D-xylose and polyphosphate catalyzed by enzyme cascades in one pot. The synthetic enzymatic pathway comprised of xylose isomerase and xylulokinase was designed to produce Xu5P, along with a third enzyme, polyphosphate kinase, responsible for in site ATP regeneration. Due to the promiscuous activity of the ATP-based xylulokinase from a hyperthermophilic bacterium Thermotoga maritima on polyphosphate, the number of enzymes in the pathway was minimized to two without polyphosphate kinase. The reactions catalyzed by the two-enzyme and three-enzyme pathways were compared for Xu5P production, and the reaction conditions were optimized by examining effects of reaction temperature, enzyme ratio and substrate concentration. The optimized two-enzyme system produced 32mM Xu5P from 50mM xylose and polyphosphate after 36 h at 45C. Biosynthesis of less costly Xu5P from D-xylose and polyphosphate could be highly feasible via this minimized two-enzyme pathway. [ABSTRACT FROM AUTHOR]

BACKGROUND The isomerization of glucose to fructose is one of the most widely used processes in the food industry to produce dietetic foods and drinks. Since this reaction is affected by external mass transfer resistances, application of an impinging jets technique in reducing external resistances may be justified. RESULTS Conversion of glucose to fructose was studied in a novel type of impinging streams reactor using glucose isomerase enzyme. Response surface methodology was applied to determine the effects of certain pertinent parameters of the process including axial velocity (A), feed concentration (B), nozzle flow rates (C) and enzyme loading (D) on the performance of the apparatus. A quadratic polynomial multiple regression model was selected and analysis of variance revealed a high coefficient of determination ( R2) value equal to 0.972. The optimum process conditions for fructose production were established as, A = 0.1 m s-1, B = 1.5 kg m-3, C = 3.3e-5 m3 s-1 and D = 0.0035 kg. CONCLUSION It was found that an impinging streams reactor had higher performance capability in comparison with conventional reactors in solid-liquid enzyme reactions. In such a reactor 30% conversion of glucose was obtained within 9 s while the same conversion in packed-bed reactors requires 5 min. © 2013 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

BACKGROUND: Simultaneous xylose isomerization and fermentation was investigated to improve the lactic acid production from xylose by Lactobacillus pentosus in a novel two-in-one bioreactor constructed by packing the immobilized xylose isomerase (65 g) in a fixed bed reactor (diameter 56 mm × 66 mm, packing volume 154 mL) with a permeable wall, which was installed inside a conventional fermenter (2 L) and rotated along the axis together with the mechanical stirrer of the fermenter. RESULTS: Xylose (20 g L−1) was completely consumed within 24 h in the novel bioreactor, compared with 72 h needed for the control without packed enzyme. The maximum cell density (17.5 g L−1) in the novel bioreactor was twice that in the control and the lactic acid productivity (0.58 g L−1 h−1) was 3.8 times higher. Repeated use of the immobilized enzyme showed that the lactic acid productivity and yield obviously dropped after the first batch fermentation but maintained almost unchanged afterwards. CONCLUSION: Simultaneous xylose isomerization and fermentation significantly improved lactic acid production from xylose by Lactobacillus pentosus. The novel bioreactor made it easier to recycle and reuse the immobilized enzyme. © 2012 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

The demand for biofuel ethanol made from clean, renewable nonfood sources is growing. Cellulosic biomass, such as switch grass (Panicum virgatum L.), is an alternative feedstock for ethanol production; however, cellulosic feedstock hydrolysates contain high levels of xylose, which needs to be converted to ethanol to meet economic feasibility. In this study, the effects of xylose isomerase on cell growth and ethanol production from biomass sugars representative of switch grass were investigated using low cell density cultures. The lager yeast species Saccharomyces pastorianus was grown with immobilized xylose isomerase in the fermentation step to determine the impact of the glucose and xylose concentrations on the ethanol production rates. Ethanol production rates were improved due to xylose isomerase; however, the positive effect was not due solely to the conversion of xylose to xylulose. Xylose isomerase also has glucose isomerase activity, so to better understand the impact of the xylose isomerase on S. pastorianus, growth and ethanol production were examined in cultures provided fructose as the sole carbon. It was observed that growth and ethanol production rates were higher for the fructose cultures with xylose isomerase even in the absence of xylose. To determine whether the positive effects of xylose isomerase extended to other yeast species, a side-by-side comparison of S. pastorianus and Saccharomyces cerevisiae was conducted. These comparisons demonstrated that the xylose isomerase increased ethanol productivity for both the yeast species by increasing the glucose consumption rate. These results suggest that xylose isomerase can contribute to improved ethanol productivity, even without significant xylose conversion. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012 [ABSTRACT FROM AUTHOR]

Immobilization of a thermostable D-xylose isomerase (EC 5.3.1.5) from Thermotoga neapolitana 5068 (TNXI) on chitin beads was accomplished via a N-terminal fusion with a chitin-binding domain (CBD) from a hyperthermophilic chitinase produced by Pyrococcus furiosus (PF1233) to create a fusion protein (CBD-TNXI). The turnover numbers for glucose to fructose conversion for both unbound and immobilized CBD-TNXI were greater than the wild-type enzyme: kcat (min-1) was ∼1,000, 3,800, and 5,800 at 80°C compared to 1,140, 10,350, and 7,000 at 90°C, for the wild-type, unbound, and immobilized enzymes, respectively. These kcat values for the glucose to fructose isomerization measured are the highest reported to date for any XI at any temperature. Enzyme kinetic inactivation at 100°C, as determined from a bi-phasic inactivation model, showed that the CBD-TNXI bound to chitin had a half-life approximately three times longer than the soluble wild-type TNXI (19.9 hours vs. 6.8 hours, respectively). Surprisingly, the unbound soluble CBD-TNXI had a significantly longer half-life (56.5 hours) than the immobilized enzyme. Molecular modeling results suggest that the N-terminal fusion impacted subunit interactions, thereby contributing to the enhanced thermostability of both the unbound and immobilized CBD-TNXI. These interactions likely also played a role in modifying active site structure, thereby diminishing substrate-binding affinities and generating higher turnover rates in the unbound fusion protein. © 2010 American Institute of Chemical Engineers Biotechnol. Prog., 2010 [ABSTRACT FROM AUTHOR]

Food and feed is possibly the area where processing anchored in biological agents has the deepest roots. Despite this, process improvement or design and implementation of novel approaches has been consistently performed, and more so in recent years, where significant advances in enzyme engineering and biocatalyst design have fastened the pace of such developments. This paper aims to provide an updated and succinct overview on the applications of enzymes in the food sector, and of progresses made, namely, within the scope of tapping formore efficient biocatalysts, through screening, structuralmodification, and immobilization of enzymes. Targeted improvements aim at enzymes with enhanced thermal and operational stability, improved specific activity, modification of pH-activity profiles, and increased product specificity, among others. This has been mostly achieved through protein engineering and enzyme immobilization, along with improvements in screening. The latter has been considerably improved due to the implementation of high-throughput techniques, and due to developments in protein expression and microbial cell culture. Expanding screening to relatively unexplored environments (marine, temperature extreme environments) has also contributed to the identification and development of more efficient biocatalysts. Technological aspects are considered, but economic aspects are also briefly addressed. [ABSTRACT FROM AUTHOR]

The effects of divalent metal cations on structural thermostability and the inactivation kinetics of homologous class IId-xylose isomerases (XI; EC 5.3.1.5) from mesophilic (Escherichia coliandBacillus licheniformis), thermophilic (Thermoanaerobacterium thermosulfurigenes), and hyperthermophilic (Thermotoga neapolitana) bacteria were examined. Unlike the three less thermophilic XIs that were substantially structurally stabilized in the presence of Co2+ or Mn2+ (and Mg2+ to a lesser extent), the melting temperature[(Tm)≈100 °C] ofT. neapolitanaXI (TNXI) varied little in the presence or absence of a single type of metal. In the presence of any two of these metals, TNXI exhibited a second melting transition between 110 °C and 114 °C. TNXI kinetic inactivation, which was non-first order, could be modeled as a two-step sequential process. TNXI inactivation in the presence of 5 mmmetal at 99–100 °C was slowest in the presence of Mn2+[half-life (t1/2) of 84 min], compared to Co2+ (t1/2 of 14 min) and Mg2+ (t1/2 of 2 min). While adding Co2+ to Mg2+ increased TNXI'st1/2 at 99–100 °C from 2 to 7.5 min, TNXI showed no significant activity at temperatures above the first melting transition. The results reported here suggest that, unlike the other class II XIs examined, single metals are required for TNXI activity, but are not essential for its structural thermostability. The structural form corresponding to the second melting transition of TNXI in the presence of two metals is not known, but likely results from cooperative interactions between dissimilar metals in the two metal binding sites. [ABSTRACT FROM AUTHOR]

The first eukaryotic xylose isomerase protein was purified from barley Hordeum vulgare. The enzyme requires Mn2+ for its activity and is fairly thermostable, with the optimum temperature being 60°C. It showed maximum activity over a broad pH range (7.0-9.0). The molecular mass of the monomer was about 50000 Da based on the SDS/PAGE, and the calculated value from the cDNA-deduced polypeptide sequence was 53620 Da. A relative mass estimation of 100000 Da was obtained from the Superose 12 chromatography, suggesting that the barley enzyme is a dimer. The cloned corresponding cDNA sequence of 1710 nucleotides encoded a polypeptide of 480 amino acids. The genomic sequence of 4473 nucleotides, revealed that the isomerase gene contained 20 introns, all starting with GT and ending with AG. One large intron was located in the 5'untranslated region. The barley isomerase has an insertion of about 40 residues at its amino terminus when compared to the prokaryotic cluster (family) II isomerases; cluster (family) I and cluster (family) II isomerases vary from the former in an insertion of around 50 residues at their amino termini. Comparison of the barley protein with the prokaryotic isomerases shows that the conserved catalytic and metal binding regions are also well conserved in barley. [ABSTRACT FROM AUTHOR]

In this study, ethanol, methanol, ethyl acetate and water extracts of Rubia tinctorum L. ( Rubiaceae) were tested for antimicrobial activity by the disc diffusion method. From the present study it was found that Rubia tinctorum L. revealed antimicrobial activity against some Gram (+) and Gram (-) bacteria, yeasts, filamentous fungi and actinomycetes. Copyright © 2006 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]