Sulfide-quinone reductase from Rhodobacter capsulatus. Purification, cloning, and expression.
A sulfide-quinone oxidoreductase (SQR, EC 1.8.5.'.) has been purified to homogeneity from chromatophores of the non-sulfur purple bacterium Rhodobacter capsulatus DSM 155. It is composed of a single polypeptide with an apparent molecular mass of about 55 kDa, exhibiting absorption and fluorescence spectra typical for a flavoprotein and similar to the SQR from the cyanobacterium Oscillatoria limnetica. From N-terminal and tryptic peptide sequences of the pure protein a genomic DNA clone was obtained by polymerase chain reaction amplification. Its sequence contains an open reading frame of 1275 base pairs (EMBL nucleotide sequence data base, accession no. X97478X97478) encoding the SQR of R. capsulatus. The deduced polypeptide consists of 425 amino acid residues with a molecular mass of 47 kDa and a net charge of +9. The high similarity (72%)/identity (48%) between the N termini of the cyanobacterial and the bacterial enzyme was confirmed and extended. Both enzymes exhibit the FAD/NAD(P) binding betaalphabeta-fold (Wierenga, R. K., Terpstra, P., and Hol, W. G. S. (1986) J. Mol. Biol. 187, 101-107). The complete sequence of the SQR from R. capsulatus shows further similarity to flavoproteins, in particular glutathione reductase and lipoamide dehydrogenase. The cloned sqr was expressed in Escherichia coli in a functional form.
Schütz
M
M
Universität Regensburg, Lehrstuhl für Zellbiologie und Pflanzenphysiologie, 93040 Regensburg, Germany.
Shahak
Y
Y
Padan
E
E
Hauska
G
G
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D017124:Conserved Sequence; D004269:DNA, Bacterial / Q000737:chemistry; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism; D008969:Molecular Sequence Data; D011808:Quinone Reductases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D016354:Rhodobacter capsulatus / Q000201:enzymology*; D016415:Sequence Alignment; D013050:Spectrometry, Fluorescence
D004269:DNA, Bacterial; D005182:Flavin-Adenine Dinucleotide; D011808:Quinone Reductases; C073412:sulfide quinone reductase