Dataset Information

Cytoplasmic Localization of Sulfide:Quinone Oxidoreductase and Persulfide Dioxygenase of Cupriavidus pinatubonensis JMP134.

ABSTRACT: Heterotrophic bacteria have recently been reported to oxidize sulfide to sulfite and thiosulfate by using sulfide:quinone oxidoreductase (SQR) and persulfide dioxygenase (PDO). In chemolithotrophic bacteria, both SQR and PDO have been reported to function in the periplasmic space, with SQR as a peripheral membrane protein whose C terminus inserts into the cytoplasmic membrane and PDO as a soluble protein. Cupriavidus pinatubonensis JMP134, best known for its ability to degrade 2,4-dichlorophenoxyacetic acid and other aromatic pollutants, has a gene cluster of sqr and pdo encoding C. pinatubonensis SQR (CpSQR) and CpPDO2. When cloned in Escherichia coli, the enzymes are functional. Here we investigated whether they function in the periplasmic space or in the cytoplasm in heterotrophic bacteria. By using sequence analysis, biochemical detection, and green fluorescent protein (GFP)/PhoA fusion proteins, we found that CpSQR was located on the cytoplasmic side of the membrane and CpPDO2 was a soluble protein in the cytoplasm with a tendency to be peripherally located near the membrane. The location proximity of these proteins near the membrane in the cytoplasm may facilitate sulfide oxidation in heterotrophic bacteria. The information may guide the use of heterotrophic bacteria in bioremediation of organic pollutants as well as H2S.IMPORTANCE Sulfide (H2S, HS-, and S2-), which is common in natural gas and wastewater, causes a serious malodor at low levels and is deadly at high levels. Microbial oxidation of sulfide is a valid bioremediation method, in which chemolithotrophic bacteria that use sulfide as the energy source are often used to remove sulfide. Heterotrophic bacteria with SQR and PDO have recently been reported to oxidize sulfide to sulfite and thiosulfate. Cupriavidus pinatubonensis JMP134 has been extensively characterized for its ability to degrade organic pollutants, and it also contains SQR and PDO. This paper shows the localization of SQR and PDO inside the cytoplasm in the vicinity of the membrane. The information may provide guidance for using heterotrophic bacteria in sulfide bioremediation.

SUBMITTER: Gao R

PROVIDER: S-EPMC5691403 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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Cytoplasmic Localization of Sulfide:Quinone Oxidoreductase and Persulfide Dioxygenase of Cupriavidus pinatubonensis JMP134.

Gao Rui R Liu Honglei H Xun Luying L

Applied and environmental microbiology 20171116 23

Heterotrophic bacteria have recently been reported to oxidize sulfide to sulfite and thiosulfate by using sulfide:quinone oxidoreductase (SQR) and persulfide dioxygenase (PDO). In chemolithotrophic bacteria, both SQR and PDO have been reported to function in the periplasmic space, with SQR as a peripheral membrane protein whose C terminus inserts into the cytoplasmic membrane and PDO as a soluble protein. <i>Cupriavidus pinatubonensis</i> JMP134, best known for its ability to degrade 2,4-dichlor ...[more]

PMID: 28939597

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Project description: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.

Dataset Information

Cytoplasmic Localization of Sulfide:Quinone Oxidoreductase and Persulfide Dioxygenase of Cupriavidus pinatubonensis JMP134.

Publications

Cytoplasmic Localization of Sulfide:Quinone Oxidoreductase and Persulfide Dioxygenase of Cupriavidus pinatubonensis JMP134.

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