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Enzyme kinetics

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https://www.readbyqxmd.com/read/28538202/development-of-nanobiocatalysts-through-the-immobilization-of-pseudomonas-fluorescens-lipase-for-applications-in-efficient-kinetic-resolution-of-racemic-compounds
#1
Bharat P Dwivedee, Jayeeta Bhaumik, Shushil K Rai, Joydev K Laha, Uttam C Banerjee
The present work reports covalent immobilization of Pseudomonas fluorescens lipase (PFL) on functionalized multiwalled carbon nanotubes (MWCNTs) as a nanobiocatalyst (NBC). This nanobiocatalyst facilitates efficient kinetic resolution of (RS)-1-phenylethanol into (S)-1-phenylethanol [C=49.7%, eep=99.5%, ees=98.1% and E value=191.4]. The immobilized preparation (MWCNTs-PFL) showed ten-fold increase in activity, thermal stability upto 80 °C and recyclability (8 cycles). MWCNTs-PFL nanobioconjugate demonstrated better stability and enhanced activity compared to covalently immobilized PFL on other matrices (silver nanoparticles, gold nanoparticles and chitosan beads) used for the study...
May 11, 2017: Bioresource Technology
https://www.readbyqxmd.com/read/28538098/two-dimensional-metal-organic-framework-nanosheets-as-an-enzyme-inhibitor-modulation-of-the-%C3%AE-chymotrypsin-activity
#2
Ming Xu, Shuai Yuan, Xin-Yu Chen, Yu-Jie Chang, Gregory Steven Day, Zhi-Yuan Gu, Hong-Cai Zhou
Two-dimensional metal-organic framework (MOF) nanosheets are utilized as effective enzyme inhibitors, providing an in-spiring means to enhance the control of cellular processes as well as improve our understanding of the surface chemistry between MOFs and enzymes. In this paper, we demonstrated that the activity of α-Chymotrypsin (ChT) can be effectively inhibited with 96.9% inhibition by the 2-D Cu(bpy)2(OTf)2 nanosheets, while the other Zn2(bim)4 nanosheets show no significant inhibition effect towards ChT...
May 24, 2017: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/28535481/graphene-oxide-enzyme-hybrid-nanoflowers-for-efficient-water-soluble-dye-removal
#3
Hui Li, Jingwei Hou, Linlin Duan, Chao Ji, Yatao Zhang, Vicki Chen
High efficient enzyme immobilization on carbon based conductive supports could provide wide applications in energy and environmental science. Here, we synthesized a 3D flower-like structured self-assembly hybrid nanocomposite with copper phosphate, laccase, graphite oxide (GO) and carbon nanotubes (CNTs) via a facile one-pot strategy under mild conditions. The prepared nanocomposite exhibited very high enzyme loading and improved laccase activity. During the formation of the nanocomposite, the copper phosphate-laccase petals were interwined by CNTs, and GO nanosheets were further coated on the petal surface...
May 11, 2017: Journal of Hazardous Materials
https://www.readbyqxmd.com/read/28533941/changes-in-clinical-laboratory-parameters-and-pharmacodynamic-markers-in-response-to-blinatumomab-treatment-of-patients-with-relapsed-refractory-all
#4
Virginie Nägele, Andrea Kratzer, Gerhard Zugmaier, Chris Holland, Youssef Hijazi, Max S Topp, Nicola Gökbuget, Patrick A Baeuerle, Peter Kufer, Andreas Wolf, Matthias Klinger
BACKGROUND: Blinatumomab has shown a remission rate of 69% in an exploratory single-arm, phase II dose-escalation study in adult patients with relapsed/refractory B-precursor acute lymphoblastic leukemia (ALL). We evaluated changes in laboratory parameters and immunopharmacodynamic markers in patients who received blinatumomab in the exploratory phase II study. METHODS: Data from 36 adults with relapsed/refractory ALL receiving blinatumomab as 4-week continuous IV infusions in various dose cohorts were analyzed for changes in liver enzymes, first-dose parameters, peripheral blood cell subpopulations, and cytokine/granzyme B release...
2017: Experimental Hematology & Oncology
https://www.readbyqxmd.com/read/28531862/unravelling-the-suitability-of-biological-induction-for-halophilic-lipase-production-by-halomonas-sp-lm1c-cultures
#5
Esther Gutiérrez-Arnillas, María Arellano, Francisco J Deive, Ana Rodríguez, María Ángeles Sanromán
In this study, the viability of using biological induction as an alternative to the conventional chemical induction in lipase production by a novel halophilic microorganism, Halomonas sp. LM1C, has been demonstrated. Thus, a 9-times increase of lipase activity (3000U/L) was recorded when Staphylococcus equorum sp. AMC7 was present in the medium, which is competitive with the results obtained when Triton X-100 was added as chemical inducer. The GC-MS data allowed concluding the true nature of the biological inducer effect, as the existence of high percentages of isomeric forms of pentadecanoic acid were detected...
May 4, 2017: Bioresource Technology
https://www.readbyqxmd.com/read/28531185/thermal-stability-and-kinetic-constants-for-129-variants-of-a-family-1-glycoside-hydrolase-reveal-that-enzyme-activity-and-stability-can-be-separately-designed
#6
Dylan Alexander Carlin, Siena Hapig-Ward, Bill Wayne Chan, Natalie Damrau, Mary Riley, Ryan W Caster, Bowen Bethards, Justin B Siegel
Accurate modeling of enzyme activity and stability is an important goal of the protein engineering community. However, studies seeking to evaluate current progress are limited by small data sets of quantitative kinetic constants and thermal stability measurements. Here, we report quantitative measurements of soluble protein expression in E. coli, thermal stability, and Michaelis-Menten constants (kcat, KM, and kcat/KM) for 129 designed mutants of a glycoside hydrolase. Statistical analyses reveal that functional Tm is independent of kcat, KM, and kcat/KM in this system, illustrating that an individual mutation can modulate these functional parameters independently...
2017: PloS One
https://www.readbyqxmd.com/read/28528213/optimal-electrostatic-interactions-between-substrate-and-protein-are-essential-for-radical-chemistry-in-ornithine-4-5-aminomutase
#7
Caitlyn Makins, Douglas A Whitelaw, Michael McGregor, Alix Petit, Robert G Mothersole, Kathleen E Prosser, Kirsten R Wolthers
Ornithine 4,5-aminomutase (OAM) from Clostridium sticklandii is an adenosylcobalamin (AdoCbl) and pyridoxal 5'-phosphate (PLP)-dependent enzyme that catalyzes a 1,2-amino shift, interconverting D-ornithine and 2S, 4R-diaminopentanoate. The reaction occurs via a radical-based mechanism whereby a PLP-bound substrate radical undergoes intramolecular isomerization via an azacyclopropylcarbinyl radical intermediate. Herein, we investigated the catalytic role of active site residues that form non-covalent interactions with PLP and/or substrate, D-ornithine...
May 17, 2017: Biochimica et Biophysica Acta
https://www.readbyqxmd.com/read/28527406/the-relevance-of-ki-calculation-for-bi-substrate-enzymes-illustrated-by-kinetic-evaluation-of-a-novel-lysine-k-acetyltransferase-8-inhibitor
#8
Hannah Wapenaar, Thea van den Bosch, Niek G J Leus, Petra E van der Wouden, Nikolaos Eleftheriadis, Jos Hermans, Gebremedhin Solomon Hailu, Dante Rotili, Antonello Mai, Alexander Dömling, Rainer Bischoff, Hidde J Haisma, Frank J Dekker
Histone acetyltransferases (HATs) are important mediators of epigenetic post-translational modifications of histones that play important roles in health and disease. A disturbance of these modifications can result in disease states, such as cancer or inflammatory diseases. Inhibitors of HATs (HATi) such as lysine (K) acetyltransferase 8 (KAT8), could be used to study the epigenetic processes in diseases related to these enzymes or to investigate HATs as therapeutic targets. However, the development of HATi is challenged by the difficulties in kinetic characterization of HAT enzymes and their inhibitors to enable calculation of a reproducible inhibitory potency...
May 5, 2017: European Journal of Medicinal Chemistry
https://www.readbyqxmd.com/read/28526768/direct-comparison-of-the-enzymatic-characteristics-and-superoxide-production-of-the-four-aldehyde-oxidase-enzymes-present-in-mouse
#9
Gokhan Kucukgoze, Mineko Terao, Enrico Garattini, Silke Leimkuhler
Aldehyde oxidases (AOXs) are molybdoflavoenzymes with an important role in the metabolism and detoxification of heterocyclic compounds and aliphatic as well as aromatic aldehydes. The enzymes use oxygen as the terminal electron acceptor and produce reduced oxygen species during turnover. Four different enzymes, mAOX1, mAOX3, mAOX4 and mAOX2, which are the products of distinct genes, are present in the mouse. A direct and simultaneous comparison of the enzymatic properties and characteristics of the four enzymes has never been performed...
May 19, 2017: Drug Metabolism and Disposition: the Biological Fate of Chemicals
https://www.readbyqxmd.com/read/28522605/structure-function-analysis-of-human-sucrase-isomaltase-identifies-key-residues-required-for-catalytic-activity
#10
Birthe Gericke, Natalie Schecker, Mahdi Amiri, Hassan Y Naim
Sucrase-isomaltase (SI) is an intestinal membrane-associated α-glucosidase that breaks down di- and oligosaccharides to absorbable monosaccharides. SI has two homologous functional subunits (sucrase and isomaltase) that both belong to the glycoside hydrolase family 31 (GH31) and differ in substrate specificity. All GH31 enzymes share a consensus sequence harboring an aspartic acid residue as a catalytic nucleophile. Moreover, crystallographic structural analysis of isomaltase predicts that another aspartic acid residue functions as a proton donor in hydrolysis...
May 18, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/28522197/identification-of-the-upstream-4-chlorophenol-biodegradation-pathway-using-a-recombinant-monooxygenase-from-arthrobacter-chlorophenolicus-a6
#11
Su Yeon Cho, Oh Sung Kwean, Jun Won Yang, Wooyoun Cho, Seonyeong Kwak, Sungyoon Park, Yejee Lim, Han S Kim
This study aimed to clarify the initial 4-chlorophenol (4-CP) biodegradation pathway promoted by a two-component flavin-diffusible monooxygenase (TC-FDM) consisting of CphC-I and CphB contained in Arthrobacter chlorophenolicus A6 and the decomposition function of CphC-I. The TC-FDM genes were cloned from A. chlorophenolicus A6, and the corresponding enzymes were overexpressed. Since CphB was expressed in an insoluble form, Fre, a flavin reductase obtained from Escherichia coli, was used. These enzymes were purified using Ni(2+)-NTA resin...
May 4, 2017: Bioresource Technology
https://www.readbyqxmd.com/read/28521084/design-of-modular-protein-tags-for-the-orthogonal-covalent-bond-formation-at-specific-dna-sequences
#12
Thang Minh Nguyen, Eiji Nakata, Masayuki Saimura, Huyen Dinh, Takashi Morii
Simultaneous formation of specific covalent linkages at nucleotides in given DNA sequences demand distinct orthogonal reactivity of DNA modification agents. Such highly specific reactions require well-balanced reactivity and affinity of the DNA modification agents. Conjugation of a sequence-specific DNA binding zinc finger protein and a self-ligating protein-tag provides a modular adaptor that expedites formation of a covalent bond between the protein-tag and a substrate-modified nucleotide at a specific DNA sequence...
May 18, 2017: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/28516418/improvement-of-the-stabilization-and-activity-of-protocatechuate-3-4-dioxygenase-isolated-from-rhizobium-sp-lmb-1-and-immobilized-on-fe3o4-nanoparticles
#13
Li-Shuang Zhang, Yue Fang, Ying Zhou, Bang-Ce Ye
Protocatechuate 3,4-dioxygenase (P34O), which is isolated from Rhizobium sp. LMB-1, catalyzes the ring cleavage step in the metabolism of aromatic compounds, and has great potential for environmental bioremediation. However, its structure is very sensitive to different environmental factors, which weaken its activity. Immobilization of the enzyme can improve its stability, allow reusability, and reduce operation costs. In this work, the relative molecular mass of the native P34O enzyme was determined to be 500 kDa by gel filtration chromatography on Sephadex G-200, and the enzyme was immobilized onto (3-aminopropyl) triethoxysilane-modified Fe3O4 nanoparticles (NPs) by the glutaraldehyde method...
May 17, 2017: Applied Biochemistry and Biotechnology
https://www.readbyqxmd.com/read/28515463/enzyme-characteristics-of-pathogen-specific-trehalose-6-phosphate-phosphatases
#14
Megan Cross, Siji Rajan, Janine Chekaiban, Jake Saunders, Chloe Hamilton, Jeong-Sun Kim, Mark J Coster, Robin B Gasser, Andreas Hofmann
Owing to the key role of trehalose in pathogenic organisms, there has recently been growing interest in trehalose metabolism for therapeutic purposes. Trehalose-6-phosphate phosphatase (TPP) is a pivotal enzyme in the most prominent biosynthesis pathway (OtsAB). Here, we compare the enzyme characteristics of recombinant TPPs from five important nematode and bacterial pathogens, including three novel members of this protein family. Analysis of the kinetics of trehalose-6-phosphate hydrolysis reveals that all five enzymes display a burst-like kinetic behaviour which is characterised by a decrease of the enzymatic rate after the pre-steady state...
May 17, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28512092/quantitative-kinetic-analyses-of-shutting-off-a-two-component-system
#15
Rong Gao, Ann M Stock
Cells rely on accurate control of signaling systems to adapt to environmental perturbations. System deactivation upon stimulus removal is as important as activation of signaling pathways. The two-component system (TCS) is one of the major bacterial signaling schemes. In many TCSs, phosphatase activity of the histidine kinase (HK) is believed to play an essential role in shutting off the pathway and resetting the system to the prestimulus state. Two basic challenges are to understand the dynamic behavior of system deactivation and to quantitatively evaluate the role of phosphatase activity under natural cellular conditions...
May 16, 2017: MBio
https://www.readbyqxmd.com/read/28511864/metabolism-and-disposition-of-14-c-methycyclosiloxanes-in-rats
#16
Jeanne Y Domoradzki, Christopher M Sushynski, Jacob M Sushynski, Debra A McNett, Cynthia Van Landingham, Kathleen P Plotzke
Octamethylcyclotetrasiloxane (D4) and decamethylcyclopentasiloxane (D5) are low molecular weight cyclic volatile methyl siloxanes (cVMSs) primarily used as intermediates or monomers in the production of high molecular weight silicone polymers. The use of D4 as a direct ingredient in personal care products has declined significantly over the past 10 years, although it may be present as a residual impurity in a variety of consumer products. D5 is still used as an intentional ingredient in cosmetics, consumer products and in dry cleaning...
May 13, 2017: Toxicology Letters
https://www.readbyqxmd.com/read/28510452/rational-optimization-of-mechanism-based-inhibitors-through-determination-of-the-microscopic-rate-constants-of-inactivation
#17
Carter G Eiden, Kimberly M Maize, Barry C Finzel, John D Lipscomb, Courtney C Aldrich
Mechanism-based inhibitors (MBIs) are widely employed in chemistry, biology, and medicine because of their exquisite specificity and sustained duration of inhibition. Optimization of MBIs is complicated because of time-dependent inhibition resulting from multistep inactivation mechanisms. The global kinetic parameters kinact and KI have been used to characterize MBIs, but they provide far less information than is commonly assumed, as shown by derivation and simulation of these parameters. We illustrate an alternative and more rigorous approach for MBI characterization through determination of the individual microscopic rate constants...
May 18, 2017: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/28510212/supercoiling-biases-the-formation-of-loops-involved-in-gene-regulation
#18
REVIEW
Laura Finzi, David Dunlap
The function of DNA as a repository of genetic information is well-known. The post-genomic effort is to understand how this information-containing filament is chaperoned to manage its compaction and topological states. Indeed, the activities of enzymes that transcribe, replicate, or repair DNA are regulated to a large degree by access. Proteins that act at a distance along the filament by binding at one site and contacting another site, perhaps as part of a bigger complex, create loops that constitute topological domains and influence regulation...
November 2016: Biophysical Reviews
https://www.readbyqxmd.com/read/28509370/use-of-mushroom-tyrosinase-to-introduce-michaelis-menten-enzyme-kinetics-to-biochemistry-students
#19
William H Flurkey, Jennifer K Inlow
An inexpensive enzyme kinetics laboratory exercise for undergraduate biochemistry students is described utilizing tyrosinase from white button mushrooms. The exercise can be completed in one or two three-hour lab sessions. The optimal amounts of enzyme, substrate (catechol), and inhibitor (kojic acid) are first determined, and then kinetic data is collected in the absence and presence of the inhibitor. A Microsoft Excel template is used to plot the data and to fit the Michaelis-Menten equation to the data to determine the kinetic parameters Vmax and Km ...
May 2017: Biochemistry and Molecular Biology Education
https://www.readbyqxmd.com/read/28505361/the-possible-evolution-and-future-of-co2-concentrating-mechanisms
#20
John A Raven, John Beardall, Patricia Sánchez-Baracaldo
CO2-concentrating mechanisms (CCMs), based either on active transport of inorganic carbon (biophysical CCMs) or on biochemistry involving supplementary carbon fixation into C4 acids (C4 and CAM), play a major role in global primary productivity. However, the ubiquitous CO2-fixing enzyme in autotrophs, Rubisco, evolved at a time when atmospheric CO2 levels were very much higher than today and O2 was very low and, as CO2 and O2 approached (by no means monotonically), today's levels, at some time subsequently many organisms evolved a CCM that increased the supply of CO2 and decreased Rubisco oxygenase activity...
May 13, 2017: Journal of Experimental Botany
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