Mechanisms of Viscous Media Effects on Elementary Steps of Bacterial Bioluminescent Reaction
Enzymes activity in a cell is determined by many factors, among which viscosity of the microenvironment plays a significant role. Various cosolvents can imitate intracellular conditions in vitro, allowing to reduce a combination of different regulatory effects. The aim of the study was to analyze th...
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2021-08-01
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| Series: | International Journal of Molecular Sciences |
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| author | Albert E. Lisitsa Lev A. Sukovatyi Sergey I. Bartsev Anna A. Deeva Valentina A. Kratasyuk Elena V. Nemtseva |
| author_facet | Albert E. Lisitsa Lev A. Sukovatyi Sergey I. Bartsev Anna A. Deeva Valentina A. Kratasyuk Elena V. Nemtseva |
| author_sort | Albert E. Lisitsa |
| collection | DOAJ |
| description | Enzymes activity in a cell is determined by many factors, among which viscosity of the microenvironment plays a significant role. Various cosolvents can imitate intracellular conditions in vitro, allowing to reduce a combination of different regulatory effects. The aim of the study was to analyze the media viscosity effects on the rate constants of the separate stages of the bacterial bioluminescent reaction. Non-steady-state reaction kinetics in glycerol and sucrose solutions was measured by stopped-flow technique and analyzed with a mathematical model developed in accordance with the sequence of reaction stages. Molecular dynamics methods were applied to reveal the effects of cosolvents on luciferase structure. We observed both in glycerol and in sucrose media that the stages of luciferase binding with flavin and aldehyde, in contrast to oxygen, are diffusion-limited. Moreover, unlike glycerol, sucrose solutions enhanced the rate of an electronically excited intermediate formation. The MD simulations showed that, in comparison with sucrose, glycerol molecules could penetrate the active-site gorge, but sucrose solutions caused a conformational change of functionally important <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><mi>α</mi></semantics></math></inline-formula>Glu175 of luciferase. Therefore, both cosolvents induce diffusion limitation of substrates binding. However, in sucrose media, increasing enzyme catalytic constant neutralizes viscosity effects. The activating effect of sucrose can be attributed to its exclusion from the catalytic gorge of luciferase and promotion of the formation of the active site structure favorable for the catalysis. |
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| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-10T08:45:00Z |
| publishDate | 2021-08-01 |
| publisher | MDPI AG |
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| series | International Journal of Molecular Sciences |
| spelling | doaj.art-9fe2a150e6644c5896c64fff8457ebd52023-11-22T08:01:13ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672021-08-012216882710.3390/ijms22168827Mechanisms of Viscous Media Effects on Elementary Steps of Bacterial Bioluminescent ReactionAlbert E. Lisitsa0Lev A. Sukovatyi1Sergey I. Bartsev2Anna A. Deeva3Valentina A. Kratasyuk4Elena V. Nemtseva5Biophysics Department, Siberian Federal University, Svobodny 79, 660041 Krasnoyarsk, RussiaBiophysics Department, Siberian Federal University, Svobodny 79, 660041 Krasnoyarsk, RussiaBiophysics Department, Siberian Federal University, Svobodny 79, 660041 Krasnoyarsk, RussiaBiophysics Department, Siberian Federal University, Svobodny 79, 660041 Krasnoyarsk, RussiaBiophysics Department, Siberian Federal University, Svobodny 79, 660041 Krasnoyarsk, RussiaBiophysics Department, Siberian Federal University, Svobodny 79, 660041 Krasnoyarsk, RussiaEnzymes activity in a cell is determined by many factors, among which viscosity of the microenvironment plays a significant role. Various cosolvents can imitate intracellular conditions in vitro, allowing to reduce a combination of different regulatory effects. The aim of the study was to analyze the media viscosity effects on the rate constants of the separate stages of the bacterial bioluminescent reaction. Non-steady-state reaction kinetics in glycerol and sucrose solutions was measured by stopped-flow technique and analyzed with a mathematical model developed in accordance with the sequence of reaction stages. Molecular dynamics methods were applied to reveal the effects of cosolvents on luciferase structure. We observed both in glycerol and in sucrose media that the stages of luciferase binding with flavin and aldehyde, in contrast to oxygen, are diffusion-limited. Moreover, unlike glycerol, sucrose solutions enhanced the rate of an electronically excited intermediate formation. The MD simulations showed that, in comparison with sucrose, glycerol molecules could penetrate the active-site gorge, but sucrose solutions caused a conformational change of functionally important <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><mi>α</mi></semantics></math></inline-formula>Glu175 of luciferase. Therefore, both cosolvents induce diffusion limitation of substrates binding. However, in sucrose media, increasing enzyme catalytic constant neutralizes viscosity effects. The activating effect of sucrose can be attributed to its exclusion from the catalytic gorge of luciferase and promotion of the formation of the active site structure favorable for the catalysis.https://www.mdpi.com/1422-0067/22/16/8827bacterial luciferasenon-steady-state reaction kineticsviscositydiffusion limitation |
| spellingShingle | Albert E. Lisitsa Lev A. Sukovatyi Sergey I. Bartsev Anna A. Deeva Valentina A. Kratasyuk Elena V. Nemtseva Mechanisms of Viscous Media Effects on Elementary Steps of Bacterial Bioluminescent Reaction International Journal of Molecular Sciences bacterial luciferase non-steady-state reaction kinetics viscosity diffusion limitation |
| title | Mechanisms of Viscous Media Effects on Elementary Steps of Bacterial Bioluminescent Reaction |
| title_full | Mechanisms of Viscous Media Effects on Elementary Steps of Bacterial Bioluminescent Reaction |
| title_fullStr | Mechanisms of Viscous Media Effects on Elementary Steps of Bacterial Bioluminescent Reaction |
| title_full_unstemmed | Mechanisms of Viscous Media Effects on Elementary Steps of Bacterial Bioluminescent Reaction |
| title_short | Mechanisms of Viscous Media Effects on Elementary Steps of Bacterial Bioluminescent Reaction |
| title_sort | mechanisms of viscous media effects on elementary steps of bacterial bioluminescent reaction |
| topic | bacterial luciferase non-steady-state reaction kinetics viscosity diffusion limitation |
| url | https://www.mdpi.com/1422-0067/22/16/8827 |
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