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Quinone binding site in a type VI sulfide:quinone oxidoreductase
ABSTRACT: Monotopic membrane-bound flavoproteins, sulfide:quinone oxidoreductases (SQRs), have a variety of physiological functions, including sulfide detoxification. SQR enzymes are classified into six groups. SQRs use the flavin adenine dinucleotide (FAD) cofactor to transfer electrons from sulfid...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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Springer Berlin Heidelberg
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9666304/ https://www.ncbi.nlm.nih.gov/pubmed/36219222 http://dx.doi.org/10.1007/s00253-022-12202-8 |
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author | Miklovics, Nikolett Duzs, Ágnes Balogh, Fanni Paragi, Gábor Rákhely, Gábor Tóth, András |
author_facet | Miklovics, Nikolett Duzs, Ágnes Balogh, Fanni Paragi, Gábor Rákhely, Gábor Tóth, András |
author_sort | Miklovics, Nikolett |
collection | PubMed |
description | ABSTRACT: Monotopic membrane-bound flavoproteins, sulfide:quinone oxidoreductases (SQRs), have a variety of physiological functions, including sulfide detoxification. SQR enzymes are classified into six groups. SQRs use the flavin adenine dinucleotide (FAD) cofactor to transfer electrons from sulfide to quinone. A type VI SQR of the photosynthetic purple sulfur bacterium, Thiocapsa roseopersicina (TrSqrF), has been previously characterized, and the mechanism of sulfide oxidation has been proposed. This paper reports the characterization of quinone binding site (QBS) of TrSqrF composed of conserved aromatic and apolar amino acids. Val331, Ile333, and Phe366 were identified near the benzoquinone ring of enzyme-bound decylubiquinone (dUQ) using the TrSqrF homology model. In silico analysis revealed that Val331 and Ile333 alternately connected with the quinone head group via hydrogen bonds, and Phe366 and Trp369 bound the quinones via hydrophobic interactions. TrSqrF variants containing alanine (V331A, I333A, F366A) and aromatic amino acid (V331F, I333F, F366Y), as well as a C-terminal α-helix deletion (CTD) mutant were generated. These amino acids are critical for quinone binding and, thus, catalysis. Spectroscopic analyses proved that all mutants contained FAD. I333F replacement resulted in the lack of the charge transfer complex. In summary, the interactions described above maintain the quinone molecule’s head in an optimal position for direct electron transfer from FAD. Surprisingly, the CTD mutant retained a relatively high level of specific activity while remaining membrane-anchored. This is a unique study because it focuses on the QBS and the oxidative stage of a type VI sulfide-dependent quinone reduction. KEY POINTS: • V331, I333, F366, and W369 were shown to interact with decylubiquinone in T. roseopersicina SqrF • These amino acids are involved in proper positioning of quinones next to FAD • I333 is essential in formation of a charge transfer complex from FAD to quinone GRAPHICAL ABSTRACT: [Image: see text] SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00253-022-12202-8. |
format | Online Article Text |
id | pubmed-9666304 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | Springer Berlin Heidelberg |
record_format | MEDLINE/PubMed |
spelling | pubmed-96663042022-11-17 Quinone binding site in a type VI sulfide:quinone oxidoreductase Miklovics, Nikolett Duzs, Ágnes Balogh, Fanni Paragi, Gábor Rákhely, Gábor Tóth, András Appl Microbiol Biotechnol Biotechnologically Relevant Enzymes and Proteins ABSTRACT: Monotopic membrane-bound flavoproteins, sulfide:quinone oxidoreductases (SQRs), have a variety of physiological functions, including sulfide detoxification. SQR enzymes are classified into six groups. SQRs use the flavin adenine dinucleotide (FAD) cofactor to transfer electrons from sulfide to quinone. A type VI SQR of the photosynthetic purple sulfur bacterium, Thiocapsa roseopersicina (TrSqrF), has been previously characterized, and the mechanism of sulfide oxidation has been proposed. This paper reports the characterization of quinone binding site (QBS) of TrSqrF composed of conserved aromatic and apolar amino acids. Val331, Ile333, and Phe366 were identified near the benzoquinone ring of enzyme-bound decylubiquinone (dUQ) using the TrSqrF homology model. In silico analysis revealed that Val331 and Ile333 alternately connected with the quinone head group via hydrogen bonds, and Phe366 and Trp369 bound the quinones via hydrophobic interactions. TrSqrF variants containing alanine (V331A, I333A, F366A) and aromatic amino acid (V331F, I333F, F366Y), as well as a C-terminal α-helix deletion (CTD) mutant were generated. These amino acids are critical for quinone binding and, thus, catalysis. Spectroscopic analyses proved that all mutants contained FAD. I333F replacement resulted in the lack of the charge transfer complex. In summary, the interactions described above maintain the quinone molecule’s head in an optimal position for direct electron transfer from FAD. Surprisingly, the CTD mutant retained a relatively high level of specific activity while remaining membrane-anchored. This is a unique study because it focuses on the QBS and the oxidative stage of a type VI sulfide-dependent quinone reduction. KEY POINTS: • V331, I333, F366, and W369 were shown to interact with decylubiquinone in T. roseopersicina SqrF • These amino acids are involved in proper positioning of quinones next to FAD • I333 is essential in formation of a charge transfer complex from FAD to quinone GRAPHICAL ABSTRACT: [Image: see text] SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00253-022-12202-8. Springer Berlin Heidelberg 2022-10-11 2022 /pmc/articles/PMC9666304/ /pubmed/36219222 http://dx.doi.org/10.1007/s00253-022-12202-8 Text en © The Author(s) 2022 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) . |
spellingShingle | Biotechnologically Relevant Enzymes and Proteins Miklovics, Nikolett Duzs, Ágnes Balogh, Fanni Paragi, Gábor Rákhely, Gábor Tóth, András Quinone binding site in a type VI sulfide:quinone oxidoreductase |
title | Quinone binding site in a type VI sulfide:quinone oxidoreductase |
title_full | Quinone binding site in a type VI sulfide:quinone oxidoreductase |
title_fullStr | Quinone binding site in a type VI sulfide:quinone oxidoreductase |
title_full_unstemmed | Quinone binding site in a type VI sulfide:quinone oxidoreductase |
title_short | Quinone binding site in a type VI sulfide:quinone oxidoreductase |
title_sort | quinone binding site in a type vi sulfide:quinone oxidoreductase |
topic | Biotechnologically Relevant Enzymes and Proteins |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9666304/ https://www.ncbi.nlm.nih.gov/pubmed/36219222 http://dx.doi.org/10.1007/s00253-022-12202-8 |
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