12 results on '"Enzyme kinetics"'
Search Results
2. Model for metabolic resistance against ALS inhibitors
- Author
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Richter, Otto, Langemann, Dirk, and Beffa, Roland
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ALS inhibitors ,branched chain amino acid biosynthesis ,enzyme kinetics ,metabolic resistance ,metabolic network ,Agriculture ,Botany ,QK1-989 - Abstract
Due to herbicide selection pressure metabolic resistance has evolved in many weed species. In this paper we analyse the interaction between the branched chain amino acid (BBC) pathway and detoxifying pathways for herbicide breakdown. The four phase detoxification pathway of herbicides comprising the action of P450, GST, glycosyltransferase and ABC transporter is modelled by a system of coupled enzyme kinetic reactions represented by nonlinear differential equations. The herbicide under consideration inhibits the enzyme ALS, which is the key enzyme for the biosynthesis of branched amino acids. For the kinetics of ALS a Monod approach is employed with a binding site for the inhibitor. Synthetic and detoxification pathways are coupled. The model is used to study the production of branched amino acids under the action of ALS inhibitors for different structures and modes of action of the detoxification pathway. The model is capable of generating typical dose response curves and their shift in dependence of the activity pattern of the enzymes of the detoxification pathway of the inhibitor.
- Published
- 2014
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- View/download PDF
3. 24. Steinheimer Gespräche: RNA‐Vakzine und ‐Medikamente.
- Subjects
CHEMICAL equilibrium ,ENZYME kinetics ,SURFACE interactions ,STRUCTURAL dynamics ,CHEMICAL industry - Abstract
Copyright of Nachrichten aus der Chemie 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.)
- Published
- 2022
- Full Text
- View/download PDF
4. The enigmatic conservation of enzyme dynamics in evolution
- Author
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Amnon Kohen
- Subjects
Stereochemistry ,Evolution ,Dihydrofolate reductase ,010402 general chemistry ,01 natural sciences ,Reaction rate constant ,Computational chemistry ,Kinetic isotope effect ,Enzyme kinetics ,Isotope effects ,lcsh:Science ,lcsh:Science (General) ,chemistry.chemical_classification ,biology ,010405 organic chemistry ,Limiting ,Evolutionary pressure ,0104 chemical sciences ,Enzyme ,Order (biology) ,chemistry ,biology.protein ,lcsh:Q ,Tunnelling ready state ,lcsh:Q1-390 - Abstract
Summary Examination of the chemical step catalysed by dihydrofolate reductase (DHFR) suggested preservation of an “ideal” transition state as the enzyme evolves from bacteria to human. This observation is enigmatic: since evolutionary pressure is most effective on enzymes’ second order rate constant (kcat/KM) and since the chemistry is not rate limiting on that kinetic parameter, why is the nature of the chemical step preserved? Studies addressing this question were presented in the 2015 Beilstein ESCEC Symposium and are summarized below.
- Published
- 2016
5. Biological Chemistry / Activation and activity of glycosylated KLKs 3, 4 and 11
- Author
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Guo, Shihui, Briza, Peter, Magdolen, Viktor, Brandstetter, Hans, and Goettig, Peter
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carbohydrates (lipids) ,N-linked glycosylation ,prostate cancer biomarker ,enzyme kinetics ,zymogen activation ,eukaryotic expression - Abstract
Human kallikrein-related peptidases 3, 4, 11, and KLK2, the activator of KLK3/PSA, belong to the prostatic group of the KLKs, whose major physiological function is semen liquefaction during the fertilization process. Notably, these KLKs are upregulated in prostate cancer and are used as clinical biomarkers or have been proposed as therapeutic targets. However, this potential awaits a detailed characterization of these proteases. In order to study glycosylated prostatic KLKs resembling the natural proteases, we used Leishmania (LEXSY) and HEK293 cells for secretory expression. Both systems allowed the subsequent purification of soluble pro-KLK zymogens with correct propeptides and of the mature forms. Periodic acid-Schiff reaction, enzymatic deglycosylation assays, and mass spectrometry confirmed the glycosylation of these KLKs. Activation of glycosylated pro-KLKs 4 and 11 turned out to be most efficient by glycosylated KLK2 and KLK4, respectively. By comparing the glycosylated prostatic KLKs with their non-glycosylated counterparts from Escherichia coli, it was observed that the N-glycans stabilize the KLK proteases and change their activation profiles and their enzymatic activity to some extent. The functional role of glycosylation in prostate-specific KLKs could pave the way to a deeper understanding of their biology and to medical applications. I631-B11 W_01213 (VLID)3600376
- Published
- 2018
6. Untersuchungen über den Abbaumechanismus einer gereinigten Polygalakturonase aus <em>Aspergillus niger</em>.
- Author
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Koller, A. and Neukom, H.
- Subjects
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ASPERGILLUS niger , *POLYGALACTURONASE , *ESTERIFICATION , *ENZYMES , *ENZYME kinetics , *BINDING sites - Abstract
1. Using an Aspergillus niger enzyme preparation, a pure polygalacturonase has been isolated by chromatography on cellulose phosphate. 2. The alkali-labile enzyme has an isoelectric point of 4.5 and a p11 optimum of 5.5; its activation energy is 8.3 kcal/mole. 3. The enzyme hydrolyzes 48% of the glycosidic bonds in pectic acid. Besides di- and mono-galacturonic acid, the hydrolysate mostly contains trigalacturonic acid. 4. Depending on the degree of esterification, the methyl ester of pectic acid is degraded with varying intensity. If the degree of esterification exceeds 75%, pectin is no longer degraded. Acetylation of the secondary alcohol groups, up to 70%, has little effect. 5. Di- and trigalacturonic acid are not hydrolyzed. Tetragalacturonic acid is split into mono- and trigalacturonic acid, penta- and hexagalacturonic acid into tri-, di- and monogalacturonic acid. 6. The isolated enzyme is an endopolygalacturonase. The study of the degradation products strongly suggests that two carboxyl groups of pectic acid become attached to the binding sites of the enzyme molecule. These two groups are separated from the active center by three and six galacturonic acid units respectively. 7. Blocking of the binding sites at. higher substrate concentrations causes a decrease in enzyme activity. [ABSTRACT FROM AUTHOR]
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- 1969
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7. Untersuchungen über die Reaktionsgeschwindigkeit der enzymatischen Angiotensinbildung in vitro in Abhängigkeit von der elektrolytischen Zusammensetzung des Inkubationsmediums.
- Author
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Dahlheim, H., Weber, P., Thurau, K., Herold, Ines, and Herold, Fräulein
- Published
- 1969
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- View/download PDF
8. Anreicherung und Charakterisierung der Erythrocytenangiotensinase.
- Author
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Dahlheim, H., Petschauer, K., and Thurau, K.
- Abstract
Copyright of Pflügers Archiv: European Journal of Physiology is the property of Springer Nature 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.)
- Published
- 1969
- Full Text
- View/download PDF
9. Bestimmung der Parameter Km und Vmax der Michaelis-Menten-Kinetik auf Basis der Versuchsplanung
- Author
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Yildirim, Ali
- Subjects
parameters ,Dewey Decimal Classification::500 | Naturwissenschaften::540 | Chemie ,Enzymkinetik ,Parameter ,ddc:540 ,Enzyme kinetics ,Parametereinschätzungsfehler ,parameter estimation error - Abstract
[no abstract]
- Published
- 2010
10. Experimentelle Analyse, Modellierung und biochemische Charakterisierung von Ein- und Zweiphasenreaktionen für die technische Biokatalyse
- Author
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Schmidt, Thomas Werner and Hartmeier, Winfried
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Benzaldehydlyase ,dynamische Optimierung ,Benzoin-Aldolase ,formate dehydrogenase ,Enzymkatalyse ,process optimisation ,modelling ,Ingenieurwissenschaften ,Enzymkinetik ,enzyme kinetics ,benzaldehyde lyase ,ddc:620 ,dynamische Modellierung ,Formiatdehydrogenase - Abstract
Reaction engineering is a key aspect in the optimisation of chemical reactions. However biocatalysis faces a set of problems while implementing the interdisciplinary methods from biology, thermodynamics, and engineering science. Four issues are of mayor concern: (1) arbitrary simplifications in kinetic modelling, (2) subjective choice of optimal reaction conditions, (3) only a rudimentary knowledge of environmental conditions on the progress curve of enzymatic reactions and (4) lacking consideration of the complex interaction of reaction and mass transfer in two-phase systems. Owing to these problems, a quantitative physical understanding of the kinetic and thermodynamic phenomena dwindles away and impairs high productivities. Thus, a systematic methodology was developed within this thesis in order to rationalise enzymatic reactions with the aim of a process optimisation. Two enzyme reactions were investigated due to their commercial interest in the fine chemical industry: Formate dehydrogenase from Candida boidinii (FDH) and benzaldehyde lyase from Pseudomonas fluorescens Biovar I (BAL). For the first time a mechanistic kinetic model for both FDH and BAL was developed. The FDH model demonstrated a superior fit to experimental data in comparison to the literature model. The model for BAL was developed on the basis of elementary reaction steps as well. As a result a mechanistic kinetic model for BAL was published for the first time and exhibited a prediction ability which was in excellent agreement with all experiments. Subsequently, the influence of the process conditions was modelled as a function of temperature, pH, ionic strength and DMF content, which served as cosolvent for the hydrophobic reactands. These thermodynamic equations were coupled with the kinetic model and the resulting combined model was tested for progress curve prediction under different conditions. Demonstrating good prediction in all experiments the combined model was successfully validated in a wide range of conditions (20-55°C, 25-400 mM ionic strength, pH 7.2-10.4, 5-45% DMF (v/v)). Since two-phase systems facilitate the conversion of hydrophobic compounds, the potential of hexane and methyl isobutyl ketone (MIBK) was investigated. Due to the interaction of enzyme reaction and mass transfer the evaluation of the intrinsic kinetics in two-phase systems are difficult. In order to overcome the problem, reaction conditions were rationally optimised. Under the optimised conditions the error due to mass transfer was successfully lowered to 2.2% and smaller and could be neglected, consequently. Initial rate analysis in the optimised two-phase systems indicated identical intrinsic model parameters and it was concluded, that the enzyme kinetics are not changed as a result of the interphase or dissolved organic solvent molecules in the aqueous phase. This argumentation was supported by four progress curve measurments in buffer-hexan and by analysis of an absolute engineering measure: the Hatta number. A model-assisted optimisation of the reaction conditions (e.g. temperature, pH value, DMF content, interfacial area) was subsequently carried out to maximise space-time-yield (STY). Remarkably, the values depend systematically on reaction time, enzyme concentration and the volume ratio of a two-phase-system. As a result, the identified optimal reaction conditions holds only true as long as the process remains unchanged. Based on the in silico-tests two optimal experiments were performed in buffer-MIBK. In comparison to literare values the productivity could be increased by 4- and 11-fold, respectively. Finally, some of these findings and additional observations associated with this study were discussed from a biochemical view. Three important results are noted: (1) For an efficient synthesis with BAL the lowest possible ionic strength should be chosen. (2) The pH dependency of the enzymatic activity showed to depend on the DMSO concentration. As a result, different pH maxima are observed for varying DMSO concentrations. Now, the seemingly contrary pH maxima (3 units difference) could be elucidated by considering this correlation in respect to the activity of BAL. (3) In this respect the claim could be supported that glutamate in position 50 governs the acidic limb of the pH-activity profile by potentiometric tests.
- Published
- 2008
11. Enzymkinetik von Phospholipase C und Aggregationsverhalten von Gentransfer-Komplexen
- Author
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Galneder, Reinhard Josef, Sackmann, Erich (Prof. Dr.), Dietrich, Klaus (Prof. Dr. Dr. h. c.), and Schliwa, Manfred (Prof. Dr.)
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enzyme kinetics ,phospholipase C ,laser tweezer ,microelectrophoresis ,zetapotential ,phosphatidylinositol-4 ,5-bisphosphate ,aggregation behaviour ,gene transfer complexes ,polylysin ,quantitative fluorescence microscopy ,transfection ,cystic fibrosis ,Enzymkinetik ,Phospholipase C ,Laserfalle ,Mikroelektrophorese ,Zetapotential ,Phosphatidylinositol-4 ,5-bisphosphat ,Aggregationsverhalten ,Gentransfer-Komplexe ,Poly-L-Lysin ,Quantitative Fluoreszenzmikroskopie ,Transfektion ,Mukoviszidose ,ddc:540 ,Chemie - Abstract
Im Rahmen dieser Arbeit wurde ein Experiment zur Laserfallen-kontrollierten Mikroelektrophorese aufgebaut und getestet, das zeitaufgelöste Messungen des Oberflächenpotentials an einzelnen kolloidalen Teilchen ermöglichte. Mit dieser Methode konnte im folgenden die Enzymkinetik von Phospholipase C mit einer bisher nicht erreichten Zeitauflösung von ca. 1 sec gemessen werden. Dazu wurden Silika-Kugeln mit einem Radius von 500 nm mit einer Lipidmembran aus neutralem Phosphatidylcholin und dem negativ geladenen Substrat Phosphatidylinositol 4,5-bisphosphat (2%) beschichtet und die Änderungen des Zetapotentials durch eine kleine Zahl von membran- gebundenen Enzymen als Funktion der Zeit aufgenommen. Die eingesetzten Konzentrationen von Phospholipase C lagen dabei im nanomolaren Bereich. Des weiteren wurde das Aggregationsverhalten von DNA/Polylysin- Komplexen mit quantitativer Fluoreszenzmikroskopie untersucht. Aus der gemessenen Größenverteilung konnte die Zahl der Plasmide pro Komplex bestimmt werden. Die Verteilungsfunktion der kolloidalen Aggregation von DNA/Polylysin-Komplexen zeigte dabei ein dynamisches Skalenverhalten. Außerdem wurde ein signifikanter Effekt von Lungen-Surfactant (Alveofact) auf das Aggregationsverhalten der Komplexe beobachtet. Die Messungen zur inneren Stabilität und Packungsdichte der Komplexe erfolgten mit der Methode des Fluoreszenz-Resonanz-Energie-Transfers in einem Spektrometer. Damit konnte die Kondensation der DNA und die Dissoziation von Komplexen durch monovalente Ionen und anionische Polymere bestimmt werden. Microelectrophoresis was combined with laser trap technology to monitor the surface potential of single colloidal particles as a function of time. This method was employed to measure the enzyme kinetics of phospholipase C (PLC) with a time resolution of approximately 1 sec. Silica-beads with 500 nm radius were coated with a phospholipid bilayer composed of electrically neutral phosphatidylcholine and the negatively charged substrate phosphatidylinositol 4,5-bisphosphate (2%). Changes in the zetapotential are caused by a few membrane bound enzymes. The coated beads were exposed to different PLC- concentrations in the nanomolar regime. Besides, quantitative fluorescence microscopy was applied to investigate the aggregation behaviour of DNA/polylysine complexes under different conditions. The number of plasmids per complex was determined with image processing by analyzing the intensity distribution of fluorescently labelled complexes. The distribution function of the colloidal aggregation process showed dynamic scaling. In the presence of lung surfactant (alveofact) a significant effect on the aggregation behaviour of DNA/polylysine complexes was observed. Furthermore, fluorescence resonance energy transfer was used to study the stability and packing density of DNA/polylysine complexes under the influence of monovalent ions and anionic polymers.
- Published
- 2007
12. Aufnahme von Cobalt, Blei und Cadmium durch Bäckerhefe
- Author
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R. Heldwein, H. W. Tromballa, and Engelbert Broda
- Subjects
Absorption (pharmacology) ,Cadmium ,Chromatography ,biology ,Chemistry ,Saccharomyces cerevisiae ,Radiochemistry ,chemistry.chemical_element ,General Medicine ,biology.organism_classification ,Applied Microbiology and Biotechnology ,Yeast ,Cell wall ,Membrane ,Biochemistry ,Toxicity ,Genetics ,ÖFOS 2012 -- NATURAL SCIENCES (1) -- Chemistry (104) -- Chemistry (1040) -- Polymer chemistry (104018) ,Enzyme kinetics ,Efflux ,Cobalt ,ÖFOS 2012 -- NATURWISSENSCHAFTEN (1) -- Chemie (104) -- Chemie (1040) -- Polymerchemie (104018) - Abstract
The toxicity for and the uptake by Saccharomyces cerevisiae of the essential trace element Co2+ and the non essential elements Cd2+ and Pb2+ were compared. Inhibition of yeast growth is observed 4 hrs after addition of Co at concentrations higher than 10−4 M. Cd is about 100 times more toxic than Co and inhibits growth immediately after addition. No toxicity of Pb is observed with concentrations up to 5 · 10−4 M. For uptake experiments the radioactive isotopes 60Co, 115mCd and 210Pb were used. The yeast samples were washed with carrier solution in order to remove the isotopes adsorbed only by the cell wall and measured by γ-scintillation counting. Co uptake by yeast is strongly dependent on glucose both under aerobic and anaerobic conditions. There is no efflux of Co once taken up. Dead yeast does not accumulate Co at all. Co already taken up is released from cells when killed, indicating that no irreversible binding in the cell interior takes place. Co transport follows biphasic saturation kinetics with the MICHAELIS constants K1 = 10−4 and K2 = 8 · 10−4 M and the maximal velocities ν1 = 9.5 and ν2 = 63 μ moles/g dry weight · hr. The Q10 for Co uptake from 10−4 M solution is 2.3. Uptake of Cd resembles that of Co. The absorption of Cd is also glucose-dependent, but independent of the presence of air, no Cd is taken up by dead cells, and Cd previously taken up is released during killing the cells. There is a considerable Pb uptake irrespective of the presence of glucose, and even by dead yeast. Killing of the yeast during uptake does not lead to loss of Pb. It is concluded that the essential trace element Co is accumulated by an energy-dependent, probably active, transport system in the membrane. Cd is similarly transported, probably because of its chemical similarity to Zn. In contrast, Pb appears to be taken up only by diffusion, but subsequently to be trapped by binding in the cell interior. In this way, the accumulation of Pb is similar in amount to those of Co or Cd although the mechanisms differ widely.
- Published
- 1977
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