Biochemical characterization of individual components of the Allochromatium vinosum DsrMKJOP transmembrane complex aids understanding of complex function in vivo.
The DsrMKJOP transmembrane complex has a most important function in dissimilatory sulfur metabolism and consists of cytoplasmic, periplasmic, and membrane integral proteins carrying FeS centers and b- and c-type cytochromes as cofactors. In this study, the complex was isolated from the purple sulfur bacterium Allochromatium vinosum and individual components were characterized as recombinant proteins. The two integral membrane proteins DsrM and DsrP were successfully produced in Escherichia coli C43(DE3) and C41(DE3), respectively. DsrM was identified as a diheme cytochrome b, and the two hemes were found to be in low-spin state. Their midpoint redox potentials were determined to be +60 and +110 mV. Although no hemes were predicted for DsrP, it was also clearly identified as a b-type cytochrome. To the best of our knowledge, this is the first time that heme binding has been experimentally proven for a member of the NrfD protein family. Both cytochromes were partly reduced after addition of a menaquinol analogue, suggesting interaction with quinones in vivo. DsrO and DsrK were both experimentally proven to be FeS-containing proteins. In addition, DsrK was shown to be membrane associated, and we propose a monotopic membrane anchoring for this protein. Coelution assays provide support for the proposed interaction of DsrK with the soluble cytoplasmic protein DsrC, which might be its substrate. A model for the function of DsrMKJOP in the purple sulfur bacterium A. vinosum is presented.
Grein
Fabian
F
Institut für Mikrobiologie & Biotechnologie, Rheinische Friedrich-Wilhelms-Universiät Bonn, Meckenheimer Allee 168, D-53115 Bonn, Germany.
Pereira
Inês A C
IA
Dahl
Christiane
C
D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D002841:Chromatiaceae / Q000235:genetics / Q000378:metabolism*; D003573:Cytochrome b Group / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D010084:Oxidation-Reduction; D013455:Sulfur / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D003573:Cytochrome b Group; D013455:Sulfur