CHARACTERIZATION OF A GLUTATHIONE-DEPENDENT FORMALDEHYDE DEHYDROGENASE FROM RHODOBACTER-SPHAEROIDES

Glutathione-dependent formaldehyde dehydrogenases (GSH-FDH) represent a ubiquitous class of enzymes, found in both prokaryotes and eukaryote s. During the course of studying energy-generating pathways in the pho tosynthetic bacterium Rhodobacter sphaeroides, a gene (adhI) encoding a GSH-FDH homolog has been identified as part of an operon (adhI-cycI) that also encodes an isoform of the cytochrome c(2) family of electro n transport proteins (isocytochrome c(2)). Enzyme assays with crude Es cherichia coli extracts expressing AdhI show that this protein has the characteristic substrate preference of a GSH-FDH. Ferguson plot analy sis with zymograms suggests that the functional form of AdhI is a homo dimer of similar to 40-kDa subunits, analogous to other GSH-FDH enzyme s. These properties of AdhI were used to show that mutations which inc rease or decrease adhI expression change the specific activity of GSH- FDH in R. sphaeroides extracts, In addition, expression of the presume d adhI-cycI operon appears to be transcriptionally regulated, since th e abundance of the major adhI-specific primer extension product is inc reased by the trans-acting spd-7 mutation, which increases the level o f both isocytochrome c(2) and AdhI activity. While transcriptional lin kage of adhI and cycI could suggest a function in a common metabolic p athway, isocytochrome c(2) (periplasm) and AdhI (cytoplasm) are locali zed in separate compartments of R. sphaeroides, Potential roles for Ad hI in carbon and energy generation and the possible relationship of GS H-FDH activity to isocytochrome c(2) will be discussed based on the co mmonly accepted physiological functions of GSH-FDH enzymes in prokaryo tes and eukaryotes.