SOP for ITC-BIND experiments
SOP describing how to perform an ITC binding experiment with a MicroCal PEAQ ITC by Malvern. Note: the DOIs from the Chemotion database for the synthesis of HK and NDK are missing in this version beacuse the DOIs do not exist yet (as of 27.04.21)
SEEK ID: https://fairdomhub.org/sops/480?version=2
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SOP_ITC#4_BIND_Gre2p.pdf
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Created: 27th Apr 2021 at 09:07
Last updated: 5th May 2021 at 12:03
AttributionsVersion History
Version 2 (latest) Created 5th May 2021 at 11:52 by Gudrun Gygli
Corrected typos, included DOIs for Chemotion for NDK and HK (if applicable).
Version 1 (earliest) Created 27th Apr 2021 at 09:07 by Gudrun Gygli
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https://orcid.org/0000-0001-9119-1778Polyglot European Scientist. I thrive working in interdisciplinary environments combining the study of enzyme reactions and mechanisms with bioinformatics, molecular modelling, automated data analysis and data stewardship.
Work of the Gygli group...
Projects: Test Project (dummy), Towards Reproducible Enzyme Modeling
Web page: Not specified
Investigation 1: An experimental workflow to provide detailed information of the molecular mechanisms of enzymes is described. This workflow will help in the application of enzymes in technical processes by providing crucial parameters needed to plan, model and implement biocatalytic processes more efficiently. These parameters are homogeneity of the enzyme sample (HES), kinetic and thermodynamic parameters of enzyme kinetics and binding of reactants to enzymes. The techniques used to measure ...
An experimental workflow to provide detailed information of the molecular mechanisms of enzymes is described. This workflow will help in the application of enzymes in technical processes by providing crucial parameters needed to plan, model and implement biocatalytic processes more efficiently. These parameters are homogeneity of the enzyme sample (HES), kinetic and thermodynamic parameters of enzyme kinetics and binding of reactants to enzymes. The techniques used to measure these properties are ...
Submitter: Gudrun Gygli
Studies: DLS measurements (homogeneity of an enzyme sample), ITC binding experiments, ITC kinetic experiments (enzyme activity), Spectrophotometric Activity Measurements
Assays: Analysis of data from ITC experiments (binding), Analysis of data from ITC experiments (kinetics), Binding of HK to Gre2p (ITC-BIND), Binding of NADP+ to Gre2p in HEPES Buffer (ITC-BIND), Binding of NADP+ to Gre2p in KPi Buffer (ITC-BIND), Binding of NADP+ to Gre2p in PBS Buffer (ITC-BIND), Binding of NADPH to Gre2p in HEPES Buffer (ITC-BIND), Binding of NADPH to Gre2p in KPi Buffer (ITC-BIND), Binding of NADPH to Gre2p in PBS Buffer (ITC-BIND), Binding of NADPH to Gre2p in Tween-KPi Buffer (ITC-BIND), Binding of NDK to Gre2p (ITC-BIND), DLS measurements in 2 buffers, DLS measurements in KPi Buffer and in KPi buffer with Tween added, DLS measurements in KPi buffer with BSA added, Kinetic parameters of Gre2p, Kinetics of the reaction of NDK and NADPH with Gre2p (ITC-MIM) in HEPES ..., Kinetics of the reaction of NDK and NADPH with Gre2p (ITC-MIM) in KPi bu..., Kinetics of the reaction of NDK and NADPH with Gre2p (ITC-MIM) in PBS bu..., Kinetics of the reaction of NDK and NADPH with Gre2p (ITC-MIM) in Tween-..., Kinetics of the reaction of NDK and NADPH with Gre2p (ITC-rSIM) in 3 buf..., Selwyn test of Gre2p, Specific activity of Gre2p
Measure binding of reactants to Gre2p by ITC (isothermal titration calorimetry).
Submitter: Gudrun Gygli
Investigation: Workflow for characterization of enzymes under ...
Assays: Analysis of data from ITC experiments (binding), Binding of HK to Gre2p (ITC-BIND), Binding of NADP+ to Gre2p in HEPES Buffer (ITC-BIND), Binding of NADP+ to Gre2p in KPi Buffer (ITC-BIND), Binding of NADP+ to Gre2p in PBS Buffer (ITC-BIND), Binding of NADPH to Gre2p in HEPES Buffer (ITC-BIND), Binding of NADPH to Gre2p in KPi Buffer (ITC-BIND), Binding of NADPH to Gre2p in PBS Buffer (ITC-BIND), Binding of NADPH to Gre2p in Tween-KPi Buffer (ITC-BIND), Binding of NDK to Gre2p (ITC-BIND)
ITC binding (BIND) experiment to determine the binding parameters of NADPH to Gre2p in 100 mM KPi Buffer.
Submitter: Gudrun Gygli
Assay type: Experimental Assay Type
Technology type: Technology Type
Investigation: Workflow for characterization of enzymes under ...
Study: ITC binding experiments
ITC binding (BIND) experiment to determine the binding parameters of NADPH to Gre2p in 1x PBS Buffer.
Submitter: Gudrun Gygli
Assay type: Experimental Assay Type
Technology type: Technology Type
Investigation: Workflow for characterization of enzymes under ...
Study: ITC binding experiments
ITC binding (BIND) experiments to determine the binding parameters of NADPH to Gre2p in 100 mM HEPES Buffer.
Submitter: Gudrun Gygli
Assay type: Experimental Assay Type
Technology type: Technology Type
Investigation: Workflow for characterization of enzymes under ...
Study: ITC binding experiments
ITC binding (BIND) experiments to determine the binding parameters of NADP+ to Gre2p in 100 mM KPi Buffer.
Submitter: Gudrun Gygli
Assay type: Experimental Assay Type
Technology type: Technology Type
Investigation: Workflow for characterization of enzymes under ...
Study: ITC binding experiments
ITC binding (BIND) experiments to determine the binding parameters of NADP+ to Gre2p in 1x PBS Buffer.
Submitter: Gudrun Gygli
Assay type: Experimental Assay Type
Technology type: Technology Type
Investigation: Workflow for characterization of enzymes under ...
Study: ITC binding experiments
ITC binding (BIND) experiments to determine the binding parameters of NADP+ to Gre2p in 100 mM HEPES Buffer.
Submitter: Gudrun Gygli
Assay type: Experimental Assay Type
Technology type: Technology Type
Investigation: Workflow for characterization of enzymes under ...
Study: ITC binding experiments
ITC binding (BIND) experiments to determine the binding parameters of NDK (nitrononane-2,8-dione) to Gre2p in 100 mM KPi Buffer. Experiments failed due to very weak binding and poor solubility of NDK in buffer.
Submitter: Gudrun Gygli
Assay type: Experimental Assay Type
Technology type: Technology Type
Investigation: Workflow for characterization of enzymes under ...
Study: ITC binding experiments
ITC binding (BIND) experiments to determine the binding parameters of HK ((5S,8S)-anti hydroxyketone) to Gre2p in 100 mM KPi Buffer. Experiments failed due to very weak binding and poor solubility of HK in buffer.
Submitter: Gudrun Gygli
Assay type: Experimental Assay Type
Technology type: Technology Type
Investigation: Workflow for characterization of enzymes under ...
Study: ITC binding experiments
ITC binding (BIND) experiment to determine the binding parameters of NADPH to Gre2p in 100 mM KPi Buffer with 0.1% Tween-20 added.
Submitter: Gudrun Gygli
Assay type: Experimental Assay Type
Technology type: Technology Type
Investigation: Workflow for characterization of enzymes under ...
Study: ITC binding experiments
