Miklovics, Nikolett and Duzs, Ágnes and Balogh, Fanni and Paragi, Gábor and Rákhely, Gábor and Tóth, András (2022) Quinone binding site in a type VI sulfide:quinone oxidoreductase. APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 106 (22). pp. 7505-7517. ISSN 0175-7598
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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 alpha-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.
| Item Type: | Article |
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| Additional Information: | Funding Agency and Grant Number: ELKH Biological Research Center; European Social Fund [EFOP-3.6.2-16-201700010, GINOP-2.3.2-15-2016-00036]; New National Excellence Program of the Ministry for Innovation and Technology [UNKP-19-3]; State of Hungary Funding text: Open access funding provided by ELKH Biological Research Center. This work headed by GR was supported by the State of Hungary, co-financed by the European Social Fund (EFOP-3.6.2-16-201700010). GP received funding from GINOP-2.3.2-15-2016-00036. NM was supported by the UNKP-19-3 New National Excellence Program of the Ministry for Innovation and Technology. |
| Uncontrolled Keywords: | Sulfur metabolism; Disulfide reductase; Quinone reduction; Sulfide: quinone oxidoreductase (SQR); Quinone binding site; |
| Subjects: | Q Science / természettudomány > QH Natural history / természetrajz > QH301 Biology / biológia > QH3020 Biophysics / biofizika |
| SWORD Depositor: | MTMT SWORD |
| Depositing User: | MTMT SWORD |
| Date Deposited: | 17 Feb 2023 12:25 |
| Last Modified: | 17 Feb 2023 12:26 |
| URI: | http://real.mtak.hu/id/eprint/159336 |
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