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ThermoSlope: A Software for Determining Thermodynamic Parameters from Single Steady-State Experiments

The determination of the temperature dependence of enzyme catalysis has traditionally been a labourious undertaking. We have developed a new approach to the classical Arrhenius parameter estimation by fitting the change in velocity under a gradual change in temperature. The evaluation with a simulat...

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Autores principales: Lund, Bjarte Aarmo, Brandsdal, Bjørn Olav
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8658824/
https://www.ncbi.nlm.nih.gov/pubmed/34885737
http://dx.doi.org/10.3390/molecules26237155
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author Lund, Bjarte Aarmo
Brandsdal, Bjørn Olav
author_facet Lund, Bjarte Aarmo
Brandsdal, Bjørn Olav
author_sort Lund, Bjarte Aarmo
collection PubMed
description The determination of the temperature dependence of enzyme catalysis has traditionally been a labourious undertaking. We have developed a new approach to the classical Arrhenius parameter estimation by fitting the change in velocity under a gradual change in temperature. The evaluation with a simulated dataset shows that the approach is valid. The approach is demonstrated as a useful tool by characterizing the Bacillus pumilus LipA enzyme. Our results for the lipase show that the enzyme is psychrotolerant, with an activation energy of 15.3 kcal/mol for the chromogenic substrate para-nitrophenyl butyrate. Our results demonstrate that this can produce equivalent curves to the traditional approach while requiring significantly less sample, labour and time. Our method is further validated by characterizing three α-amylases from different species and habitats. The experiments with the α-amylases show that the approach works over a wide range of temperatures and clearly differentiates between psychrophilic, mesophilic and thermophilic enzymes. The methodology is released as an open-source implementation in Python, available online or used locally. This method of determining the activation parameters can make studies of the temperature dependence of enzyme catalysis more widely adapted to understand how enzymes have evolved to function in extreme environments. Moreover, the thermodynamic parameters that are estimated serve as functional validations of the empirical valence bond calculations of enzyme catalysis.
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spelling pubmed-86588242021-12-10 ThermoSlope: A Software for Determining Thermodynamic Parameters from Single Steady-State Experiments Lund, Bjarte Aarmo Brandsdal, Bjørn Olav Molecules Article The determination of the temperature dependence of enzyme catalysis has traditionally been a labourious undertaking. We have developed a new approach to the classical Arrhenius parameter estimation by fitting the change in velocity under a gradual change in temperature. The evaluation with a simulated dataset shows that the approach is valid. The approach is demonstrated as a useful tool by characterizing the Bacillus pumilus LipA enzyme. Our results for the lipase show that the enzyme is psychrotolerant, with an activation energy of 15.3 kcal/mol for the chromogenic substrate para-nitrophenyl butyrate. Our results demonstrate that this can produce equivalent curves to the traditional approach while requiring significantly less sample, labour and time. Our method is further validated by characterizing three α-amylases from different species and habitats. The experiments with the α-amylases show that the approach works over a wide range of temperatures and clearly differentiates between psychrophilic, mesophilic and thermophilic enzymes. The methodology is released as an open-source implementation in Python, available online or used locally. This method of determining the activation parameters can make studies of the temperature dependence of enzyme catalysis more widely adapted to understand how enzymes have evolved to function in extreme environments. Moreover, the thermodynamic parameters that are estimated serve as functional validations of the empirical valence bond calculations of enzyme catalysis. MDPI 2021-11-26 /pmc/articles/PMC8658824/ /pubmed/34885737 http://dx.doi.org/10.3390/molecules26237155 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Lund, Bjarte Aarmo
Brandsdal, Bjørn Olav
ThermoSlope: A Software for Determining Thermodynamic Parameters from Single Steady-State Experiments
title ThermoSlope: A Software for Determining Thermodynamic Parameters from Single Steady-State Experiments
title_full ThermoSlope: A Software for Determining Thermodynamic Parameters from Single Steady-State Experiments
title_fullStr ThermoSlope: A Software for Determining Thermodynamic Parameters from Single Steady-State Experiments
title_full_unstemmed ThermoSlope: A Software for Determining Thermodynamic Parameters from Single Steady-State Experiments
title_short ThermoSlope: A Software for Determining Thermodynamic Parameters from Single Steady-State Experiments
title_sort thermoslope: a software for determining thermodynamic parameters from single steady-state experiments
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8658824/
https://www.ncbi.nlm.nih.gov/pubmed/34885737
http://dx.doi.org/10.3390/molecules26237155
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