Showing papers on "Substrate (chemistry) published in 2011"
TL;DR: Of the phenol molecules tested, tannic acid was the single, most damaging aromatic compound that caused both deactivation and reversible loss (inhibition) of all of enzyme activities tested.
484 citations
TL;DR: Pt-Ft possesses both catalase and peroxidase activities for different substrates under different conditions, and showed differential response to pH and temperature for different reaction substrates.
370 citations
TL;DR: The effects of biochar on enzyme activities in soils are highly variable; these effects are likely associated with reactions between biochar and the target substrate and fluorometric assays are more robust to, or account for, this sorption better than the colorimetric assays used herein.
Abstract: We studied the effects of a biochar made from fast pyrolysis of switchgrass on four soil enzymes (β-glucosidase, β-N-acetylglucosaminidase, lipase, and leucine aminopeptidase) to determine if biochar would consistently modify soil biological activities. Thus, we conducted a series of enzyme assays on biochar-amended soils. Inconsistent results from enzyme assays of char-amended soils suggested that biochar had variable effects on soil enzyme activities, thus we conducted a second experiment to determine if biochar reacts predictably with either enzyme or substrate in in vitro reactions. Both colorimetric and fluorescent assays were used for β-glucosidase and β-N-acetylglucosaminidase. Seven days after biochar was added to microcosms of 3 different soils, fluorescence-based assays revealed some increased enzyme activities (up to 7-fold for one measure of β-glucosidase in a shrub-steppe soil) and some decreased activities (one-fifth of the unamended control for lipase measured in the same shrub-steppe soil), compared to non-amended soil. In an effort understand the varied effects, purified enzymes or substrates were briefly exposed to biochar and then assayed. In contrast to the soil assays, except for β-N-acetylglucosaminidase, the exposure of substrate to biochar reduced the apparent activity of the enzymes, suggesting that sorption reactions between substrate and biochar impeded enzyme function. Our findings indicate that fluorometric assays are more robust to, or account for, this sorption better than the colorimetric assays used herein. The activity of purified β-N-acetylglucosaminidase increased 50–75% following biochar exposure, suggesting a chemical enhancement of enzyme function. In some cases, biochar stimulates soil enzyme activities, to a much greater degree than soil assays would indicate, given that substrate reactivity can be impeded by biochar exposure. We conclude that the effects of biochar on enzyme activities in soils are highly variable; these effects are likely associated with reactions between biochar and the target substrate.
346 citations
TL;DR: ZIF-8 films were successfully prepared on a flexible nylon substrate with a contra-diffusion synthesis method, and gas permeation experiments indicated that the films were continuous and compact.
302 citations
TL;DR: This work reviews numerous examples of natural and synthetic complexes featuring substrate channeling that have great biotechnological potentials in metabolic engineering, multi-enzyme-mediated biocatalysis, and cell-free synthetic pathway biotransformation (SyPaB).
263 citations
TL;DR: PcGH61D, the gene product of an open reading frame in the genome of Phanerochaete chrysosporium, is an enzyme that cleaves cellulose using a metal-dependent oxidative mechanism that leads to generation of aldonic acids.
Abstract: Many fungi growing on plant biomass produce proteins currently classified as glycoside hydrolase family 61 (GH61), some of which are known to act synergistically with cellulases. In this study we show that PcGH61D, the gene product of an open reading frame in the genome of Phanerochaete chrysosporium, is an enzyme that cleaves cellulose using a metal-dependent oxidative mechanism that leads to generation of aldonic acids. The activity of this enzyme and its beneficial effect on the efficiency of classical cellulases are stimulated by the presence of electron donors. Experiments with reduced cellulose confirmed the oxidative nature of the reaction catalyzed by PcGH61D and indicated that the enzyme may be capable of penetrating into the substrate. Considering the abundance of GH61-encoding genes in fungi and genes encoding their functional bacterial homologues currently classified as carbohydrate binding modules family 33 (CBM33), this enzyme activity is likely to turn out as a major determinant of microbial biomass-degrading efficiency.
253 citations
TL;DR: In this paper, a method for fabrication of an efficient surface enhanced Raman scattering (SERS) substrate by combination of metallic nanostructures and graphene, which shows dramatic Raman enhancement and efficient adsorption of aromatic molecules.
Abstract: We report a method for fabrication of an efficient surface enhanced Raman scattering (SERS) substrate by combination of metallic nanostructures and graphene, which shows dramatic Raman enhancement and efficient adsorption of aromatic molecules. As an example, the fabricated Ag or Au nanoparticle (NP)-decorated reduced graphene oxide (rGO) on Si substrate is used as an efficient SERS substrate to detect the adsorbed aromatic molecules with a low detection limit at nM level. Systematic studies on the effects of NP size and substrate morphology on Raman enhancement are presented. This method might be useful for the future application in detection of biomolecules, such as DNA and proteins.
249 citations
TL;DR: Experimental results revealed that optimal experimental conditions can be rationally chosen to control (accelerate or restrain) this reaction, providing direct experimental evidence that surface plasmon resonance plays the most important role in these surface-catalyzed reactions.
Abstract: In this article, we experimentally investigate the substrate, wavelength, and time dependence of the plasmon-assisted surface-catalyzed dimerization of 4-nitrobenzenethiol to form p,p′-dimercaptoazobenzene on Au, Ag, and Cu films. We provide direct experimental evidence that surface plasmon resonance plays the most important role in these surface-catalyzed reactions. It is found that the reaction is strongly dependent on the substrate, the wavelength of the laser, and the reaction timescales. Our experimental results revealed that optimal experimental conditions can be rationally chosen to control (accelerate or restrain) this reaction. The experimental results are also confirmed by theoretical calculations.
226 citations
TL;DR: It is suggested that residues R209 and V218, situated in a second shell of residues surrounding the active site, play a role in substrate binding orientation based on their flexibility and position, which lends further support to a previously suggested mechanism by which monophenolic substrates dock mainly to CuA.
226 citations
TL;DR: In this paper, the cellulase enzyme was bound via physical adsorption (ionic bound), and FTIR spectroscopy confirmed the successful binding of cellulase (endoglucanase) onto the particle, and binding efficiency was determined at 95% using the Bradford method.
194 citations
TL;DR: The supramolecular tandem enzyme assay effectively measures the kinetics of thermolysin, including the accurate determination of sequence specificity, stereospecificity, endo- versus exopeptidase activity, and sensitivity to terminal charges.
Abstract: An analytical method has been developed for the continuous monitoring of protease activity on unlabeled peptides in real time by fluorescence spectroscopy. The assay is enabled by a reporter pair comprising the macrocycle cucurbit[7]uril (CB7) and the fluorescent dye acridine orange (AO). CB7 functions by selectively recognizing N-terminal phenylalanine residues as they are produced during the enzymatic cleavage of enkephalin-type peptides by the metalloendopeptidase thermolysin. The substrate peptides (e.g., Thr-Gly-Ala-Phe-Met-NH2) bind to CB7 with moderately high affinity (K ≈ 104 M–1), while their cleavage products (e.g., Phe-Met-NH2) bind very tightly (K > 106 M–1). AO signals the reaction upon its selective displacement from the macrocycle by the high affinity product of proteolysis. The resulting supramolecular tandem enzyme assay effectively measures the kinetics of thermolysin, including the accurate determination of sequence specificity (Ser and Gly instead of Ala), stereospecificity (d-Ala inst...
Patent•
09 Jun 2011
TL;DR: In this article, a method for preparing and resulting articles of manufacture comprising a substrate having a surface, a bulk beneath the surface, and a grafted polymer layer on the substrate surface, in combination, constituting a modified surface having a fibrinogen adsorption of less than about 125 ng/cm2.
Abstract: A method for preparing and resulting articles of manufacture comprising a substrate having a surface, a bulk beneath the surface, and a grafted polymer layer on the substrate surface, the substrate surface and the grafted polymer layer, in combination, constituting a modified surface having a fibrinogen adsorption of less than about 125 ng/cm2 in a fibrinogen binding assay in which the modified surface is incubated for 60 minutes at 37 C in 70 µg/mL fibrinogen derived from human plasma containing 1.4 µg/mL I-125 radiolabeled fibrinogen.
TL;DR: In this paper, the mesoporous TiO2 substrate and bifunctional linker, mercaptopropionic acid (MPA), were used to disperse and stabilize the CdS QDs.
Abstract: To accomplish the more effective coupling of cadmium sulfide quantum dots (CdS QDs), the mesoporous TiO2 substrate and bifunctional linker, mercaptopropionic acid (MPA), were used to disperse and stabilize the CdS QDs. Due to the porous nano-architecture on the TiO2 substrate with large surface area and high crystallinity, the efficiency of degradation of organic compounds in aqueous solution under visible light irradiation is greatly enhanced, compared to CdS loaded anatase TiO2 without porous structure and common commercial P25. Furthermore, the bifunctional linking molecule, MPA, could effectively disperse and stabilize CdS nanoparticles. CdS/TiO2 with the linking molecule CdS-MPA-TiO2(m) exhibits much more stability and activity than CdS-TiO2(m) which is prepared by direct deposition. After 3 cycling tests of degradation of MB (methylene blue), the loss ratio of CdS on CdS-TiO2(m) is 70.6%, much larger than that of 17.8% on CdS-MPA-TiO2(m). This work may give ideas for the synthesis of other stable and active supported catalysts in many fields.
01 Jan 2011
TL;DR: The isolated laccase can be used in development of biosensor for detecting the phenolic compounds from the effluents of paper industries and metal ions like CuSO4, BaCl2, MgCl2 and MnCl2 moderately decrease enzyme activity.
Abstract: Laccases are blue copper oxidases (E.C. 1.10.3.2 benzenediol: oxygen oxidoreductase) that catalyze the one-electron oxidation of phenolics, aromatic amines, and other electron-rich substrates with the concomitant reduction of O2 to H2O. They are currently seen as highly interesting industrial enzymes because of their broad substrate specificity. A positive strain was isolated and characterized as nonspore forming Basidiomycetes Pleurotus sp. Laccase activity was determined using ABTS as substrate. Laccase was purified by ionexchange and gel filtration chromatography. The purified laccase was a monomer showed a molecular mass of kDa as estimated by SDS-PAGE and a 72-fold purification with a 22% yield. The optimal pH and temperature were 4.5 and respectively. The and values are 250 (mM) and 0.33 (μmol/min), respectively, for ABTS as substrate. Metal ions like CuSO4, BaCl2, MgCl2, FeCl2, ZnCl2 have no effect on purified laccase whereas HgCl2 and MnCl2 moderately decrease enzyme activity. SDS and sodium azide inhibited enzyme activity, whereas Urea, PCMB, DTT, and mercaptoethanol have no effect on enzyme activity. The isolated laccase can be used in development of biosensor for detecting the phenolic compounds from the effluents of paper industries.
TL;DR: The method of immobilization can overcome the problem of reduced permeability of xylan, a high molecular weight substrate, to its enzyme which is conventionally entrapped within the alginate beads.
TL;DR: A new class of a SERS substrate with ordered nanostructures fabricated on silicon wafer using a deep UV (DUV) lithography technique followed by surface coating of silver and/or gold film is reported, which is the best among the reported SERS substrates.
TL;DR: In this paper, the effects of substrate structure on the formation of the selective layer in a two-step preparation were investigated, and the results revealed that the substrate surface structure is very important for fabricating a RO-like thin film.
TL;DR: A novel bacterial enzyme from Marinobacter aquaeolei VT8 is identified, purify, and characterize that catalyzes the reduction of fatty acyl-CoA by four electrons to the corresponding fatty alcohol, eliminating the need for a separate fatty aldehyde reductase.
Abstract: Fatty alcohols are of interest as a renewable feedstock to replace petroleum compounds used as fuels, in cosmetics, and in pharmaceuticals. One biological approach to the production of fatty alcohols involves the sequential action of two bacterial enzymes: (i) reduction of a fatty acyl-CoA to the corresponding fatty aldehyde catalyzed by a fatty acyl-CoA reductase, followed by (ii) reduction of the fatty aldehyde to the corresponding fatty alcohol catalyzed by a fatty aldehyde reductase. Here, we identify, purify, and characterize a novel bacterial enzyme from Marinobacter aquaeolei VT8 that catalyzes the reduction of fatty acyl-CoA by four electrons to the corresponding fatty alcohol, eliminating the need for a separate fatty aldehyde reductase. The enzyme is shown to reduce fatty acyl-CoAs ranging from C8:0 to C20:4 to the corresponding fatty alcohols, with the highest rate found for palmitoyl-CoA (C16:0). The dependence of the rate of reduction of palmitoyl-CoA on substrate concentration was cooperative, with an apparent K(m) ~ 4 μM, V(max) ~ 200 nmol NADP(+) min(-1) (mg protein)(-1), and n ~ 3. The enzyme also reduced a range of fatty aldehydes with decanal having the highest activity. The substrate cis-11-hexadecenal was reduced in a cooperative manner with an apparent K(m) of ~50 μM, V(max) of ~8 μmol NADP(+) min(-1) (mg protein)(-1), and n ~ 2.
TL;DR: The interaction between the binding pocket of Mv BOx and its highly conjugated natural organic substrate, bilirubin, can be used to stabilise the enzyme on a pyrolytic graphite electrode, more than doubling its electrocatalytic activity relative to the current obtained by simple adsorption of the protein to the carbon surface.
Abstract: The blue multi-copper oxidase bilirubin oxidase (BOx) from the ascomycete plant pathogen Myrothecium verrucaria (Mv) efficiently catalyses the oxidation of bilirubin to biliverdin, with the concomitant reduction of O2 to water, a reaction of considerable interest for low-temperature bio-fuel cell applications. We have solved the complete X-ray determined structure of Mv BOx at 2.4 A resolution, using molecular replacement with the Spore Coat Protein A (CotA) enzyme from Bacillus subtilis (PDB code 1GSK) as a template. The structure reveals an unusual environment around the blue type 1 copper (T1 Cu) that includes two non-coordinating hydrophilic amino acids, asparagine and threonine. The presence of a long, narrow and hydrophilic pocket near the T1 Cu suggests that structure of the substrate-binding site is dynamically determined in vivo. We show that the interaction between the binding pocket of Mv BOx and its highly conjugated natural organic substrate, bilirubin, can be used to stabilise the enzyme on a pyrolytic graphite electrode, more than doubling its electrocatalytic activity relative to the current obtained by simple adsorption of the protein to the carbon surface.
TL;DR: The noncharacterized protein ACL75304 encoded by the gene Ccel_0941 from Clostridium cellulolyticum H10 (ATCC 35319), previously proposed as the xylose isomerase domain protein TIM barrel, was cloned and expressed in Escherichia coli and characterized as d-psicose 3-epimerase.
Abstract: The noncharacterized protein ACL75304 encoded by the gene Ccel_0941 from Clostridium cellulolyticum H10 (ATCC 35319), previously proposed as the xylose isomerase domain protein TIM barrel, was cloned and expressed in Escherichia coli. The expressed enzyme was purified by nickel-affinity chromatography with electrophoretic homogeneity and then characterized as d-psicose 3-epimerase. The enzyme was strictly metal-dependent and showed a maximal activity in the presence of Co2+. The optimum pH and temperature for enzyme activity were 55 °C and pH 8.0. The half-lives for the enzyme at 60 °C were 6.8 h and 10 min when incubated with and without Co2+, respectively, suggesting that this enzyme was extremely thermostable in the presence of Co2+ but readily inactivated without metal ion. The Michaelis–Menten constant (Km), turnover number (kcat), and catalytic efficiency (kcat/Km) values of the enzyme for substrate d-psicose were estimated to be 17.4 mM, 3243.4 min–1, and 186.4 mM min–1, respectively. The enzyme ca...
TL;DR: The (R)-imine reductase (RIR) of Streptomyces sp.
Abstract: The (R)-imine reductase (RIR) of Streptomyces sp. GF3587 was purified and characterized. It was found to be a NADPH-dependent enzyme, and was found to be a homodimer consisting of 32 kDa subunits. Enzymatic reduction of 10 mM 2-methyl-1-pyrroline (2-MPN) resulted in the formation of 9.8 mM (R)-2-methylpyrrolidine ((R)-2-MP) with 99% e.e. The enzyme showed not only reduction activity for 2-MPN at neutral pH (6.5–8.0), but also oxidation activity for (R)-2-MP under alkaline pH (10–11.5) conditions. It appeared to be a sulfhydryl enzyme based on the sensitivity to sulfhydryl specific inhibitors. It was very specific to 2-MPN as substrate.
TL;DR: Investigation of the hydrolytic properties of the cellulase under different substrate concentrations at a fixed enzyme-to-substrate ratio indicated that a decline in the binding capacity of cellulase may explain the long-observed but little-understood phenomenon of a lower substrate digestibility with increased substrate concentration.
Abstract: Current technology for conversion of biomass to ethanol is an enzyme-based biochemical process. In bioethanol production, achieving high sugar yield at high solid loading in enzymatic hydrolysis step is important from both technical and economic viewpoints. Enzymatic hydrolysis of cellulosic substrates is affected by many parameters, including an unexplained behavior that the glucan digestibility of substrates by cellulase decreased under high solid loadings. A comprehensive study was conducted to investigate this phenomenon by using Spezyme CP and Avicel as model cellulase and cellulose substrate, respectively. The hydrolytic properties of the cellulase under different substrate concentrations at a fixed enzyme-to-substrate ratio were characterized. The results indicate that decreased sugar yield is neither due to the loss of enzyme activity at a high substrate concentration nor due to the higher end-product inhibition. The cellulase adsorption kinetics and isotherm studies indicated that a decline in the binding capacity of cellulase may explain the long-observed but little-understood phenomenon of a lower substrate digestibility with increased substrate concentration. The mechanism how the enzyme adsorption properties changed at high substrate concentration was also discussed in the context of exploring the improvement of the cellulase-binding capacity at high substrate loading.
TL;DR: In this article, an ionic liquid was used as reaction medium to produce biodiesel (fatty acid methyl ester, FAME) with soybean oil and methanol through transesterification by Novozym 435.
TL;DR: In the sequential batch production of galacto-oligosaccharides, biocatalyst efficiency was increased by 190% with respect to the free enzyme in solution, and 8500 g of galactsiccharides per gram of enzyme preparation were produced after 10 batches, significantly higher than the maximum attainable by using the soluble enzyme.
TL;DR: In this paper, the authors described the optimization of the esterification reaction of butyl acetate synthesis catalyzed by Candida antarctica lipase B (Novozym 435).
TL;DR: Surface-enhanced Raman scattering (SERS) measurements demonstrate that the Ag nanosheet-assembled micro-hemispheres can serve as sensitive and reproducible SERS substrates.
TL;DR: In this paper, the growth mechanisms of N-doped graphene at low and high substrate temperatures were discussed, and it was shown that high substrate temperature (TS) is the best temperature for the formation of high TS.
Abstract: Chemical doping of graphene with foreign atoms is one of the most promising ways to modify the electronic structure of graphene. We fabricated nitrogen (N)-doped graphene on a Pt(111) surface through a chemical vapor deposition method; the heated substrate was exposed to such N-containing organic molecules as pyridine and acrylonitrile. Analysis by X-ray photoelectron spectroscopy (XPS) and Raman spectroscopy revealed that N-doped graphene was formed on a Pt(111) surface from pyridine at the substrate temperature (TS) higher than 500 °C, while nitrogen was not doped at TS higher than 700 °C. Exposing the heated substrate to acrylonitrile also led to formation of graphene but nitrogen was not incorporated at any TS. On the basis of the experimental results, we discuss the growth mechanisms of N-doped graphene at low and high TS.
Patent•
12 Oct 2011TL;DR: A glass for substrate, which consists, as represented by mass percentage, essentially of: SiO2 40 to 59%, Al2O3 5 to 20%, B2O2 0 to 8, MgO 0 to 10, CaO 0, 12, SrO 2 to 20, BaO 0to 2, ZnO0 to 4, Li2O 0- 2, Na2O, K2O0, K 2O0-12, TiO2, and ZrO 2 0 to 5%, wherein SiO+CaO+Sr
Abstract: A glass for substrate, which consists, as represented by mass percentage, essentially of: SiO2 40 to 59%, Al2O3 5 to 20%, B2O3 0 to 8%, MgO 0 to 10%, CaO 0 to 12%, SrO 2 to 20%, BaO 0 to 2%, ZnO 0 to 4%, Li2O 0 to 2%, Na2O 0 to 10%, K2O 0 to 12%, TiO2 0 to 10%, and ZrO2 0 to 5%, wherein MgO+CaO+SrO+BaO is at least 15%.