Bacterial Luciferases from Vibrio harveyi and Photobacterium leiognathi Demonstrate Different Conformational Stability as Detected by Time-Resolved Fluorescence Spectroscopy
| Autor: | Dmitry V. Gulnov, Elena V. Nemtseva, Ludmila Burakova, Bogdan S. Melnik, Natalya E. Karuzina, Lev A. Sukovatyi, Valentina A. Kratasyuk, M. A. Gerasimova |
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| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
QH301-705.5
Equilibrium unfolding tryptophan fluorescence Molecular Dynamics Simulation Article Catalysis time-resolved spectroscopy Inorganic Chemistry Protein Domains Photobacterium leiognathi Denaturation (biochemistry) Luciferase Physical and Theoretical Chemistry Biology (General) Molecular Biology QD1-999 Spectroscopy Vibrio bacterial luciferase Luciferases conformational stability Quenching (fluorescence) biology Photobacterium Vibrio harveyi Chemistry Organic Chemistry General Medicine biology.organism_classification molecular dynamics Computer Science Applications Luciferases Bacterial Spectrometry Fluorescence Förster resonance energy transfer unfolding pathway Biophysics FRET urea-induced denaturation |
| Zdroj: | International Journal of Molecular Sciences Volume 22 Issue 19 International Journal of Molecular Sciences, Vol 22, Iss 10449, p 10449 (2021) |
| ISSN: | 1422-0067 |
| DOI: | 10.3390/ijms221910449 |
| Popis: | Detecting the folding/unfolding pathways of biological macromolecules is one of the urgent problems of molecular biophysics. The unfolding of bacterial luciferase from Vibrio harveyi is well-studied, unlike that of Photobacterium leiognathi, despite the fact that both of them are actively used as a reporter system. The aim of this study was to compare the conformational transitions of these luciferases from two different protein subfamilies during equilibrium unfolding with urea. Intrinsic steady-state and time-resolved fluorescence spectra and circular dichroism spectra were used to determine the stages of the protein unfolding. Molecular dynamics methods were applied to find the differences in the surroundings of tryptophans in both luciferases. We found that the unfolding pathway is the same for the studied luciferases. However, the results obtained indicate more stable tertiary and secondary structures of P. leiognathi luciferase as compared to enzyme from V. harveyi during the last stage of denaturation, including the unfolding of individual subunits. The distinctions in fluorescence of the two proteins are associated with differences in the structure of the C-terminal domain of α-subunits, which causes different quenching of tryptophan emissions. The time-resolved fluorescence technique proved to be a more effective method for studying protein unfolding than steady-state methods. |
| Databáze: | OpenAIRE |
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