Comparative characterization and expression analysis of the four Old Yellow Enzyme homologues from Shewanella oneidensis indicate differences in physiological function

Abstract

Shewanella oneidensis contains four genes that encode proteins that have high sequence identity with yeast OYE (Old Yellow Enzyme, an NADPH oxidoreductase), the well-studied archetype of the OYE protein family. The present paper describes the first comparative study of OYEs that are present in a single bacterial species, performed to gain insight into their biochemical properties and physiological importance. The four proteins [named SYE1-SYE4 (Shewanella Yellow Enzyme 1-4)] were expressed as glutathione S-transferase fusion proteins in Escherichia coli. The yield of SYE2, however, was too low for further characterization, even after expression attempts in S. oneidensis. The SYE1, SYE3 and SYE4 proteins were found to have characteristics similar to those of other OYE family members. They were identified as flavoproteins that catalyse the reduction of different alpha,beta-unsaturated carbonyl compounds and form charge transfer complexes with a range of phenolic compounds. Whereas the properties of SYE1 and SYE3 were very similar, those of SYE4 were clearly different in terms of ligand binding, catalytic efficiency and substrate specificity. Also, the activity of SYE4 was found to be NADPH-dependent, whereas SYE1 and SYE3 had a preference for NADH. It has been suggested that yeast OYE protects the actin cytoskeleton from oxidative stress. There are indications that bacterial OYEs are also involved in the oxidative stress response, but their exact role is unclear. Induction studies in S. oneidensis revealed that yeast and bacterial OYEs may share a common physiological role, i.e. the protection of cellular components against oxidative damage. As only SYE4 was induced under oxidative stress conditions, however, a functional divergence between bacterial OYEs is likely to exist.

Ligand-induced Conformational Changes in the Capping Subdomain of a Bacterial Old Yellow Enzyme Homologue and Conserved Sequence Fingerprints Provide New Insights into Substrate Binding

“…We identified four OYE homologues (NP718044, NP718043, NP719682, and NP718946) that we named SYE1-4 for Shewanella yellow enzyme (31). SYE1 (NP718044) is the Shewanella OYE homologue with the greatest sequence identity to OYE1.…”

An enoate reductase Achr-OYE4 from Achromobacter sp. JA81: characterization and application in asymmetric bioreduction of C=C bonds

“…Most characterized enoate reductases prefer NADPH over NADH as the cofactor (Chaparro-Riggers et al 2007;Toogood et al 2010), whereas only a few prefer NADH (Brige et al 2006;French and Bruce 1994), such as morphinone reductase, SYE1, and SYE3. In the present study, Achr-OYE4 displayed a preference for NADH on using all testing substrates, and the catalytic efficiency toward NADH Fig.…”

“…Achr-OYE4 contains 373 amino acid residues and is most closely related to SYE1 from Shewanella oneidensis, with 64.6 % identity (Brige et al 2006). The present study focused on the characterization of this novel enoate reductase, that is, Achr-OYE4, as well as its application in the asymmetric bioreduction of activated C=C bonds.…”

Clostridium scindens baiCD and baiH genes encode stereo-specific 7α/7β-hydroxy-3-oxo-Δ4-cholenoic acid oxidoreductases☆

“…cerevisiae, sye1 from She. oneidensis, and the yqjM gene from B. subtilis were shown to be inducible by small α/β-unsaturated aldehydes during oxidative stress responses to lipid peroxidation (47,62,64) and actin cytoskeletal oxidative damage in yeast (65). Detoxification does not appear to be the function of the bile acid 7α/β-dehydroxylating pathways, but rather the substrates appear to serve as electron sinks to enhance fermentation (2,14).…”

“…The noxB-2 gene shares 98.9 % identity with the noxB-1 gene from A. fulgidus and is found upstream of mediumchain acyl-CoA ligase in this bacterium suggesting these genes are also involved in fatty acid metabolism (55). The more distantly related Old Yellow Enzyme (OYE) family homologues are found both in eukaryotes such as Saccaromyces cerevisiae (OYE1, OYE2) (56) and S. carlsbergensis (OYE1) (60), as well as the prokaryotes including Bacillus subtilis (YqjM) (61), and Shewanella oneidensis (SYE1-SYE4) (62). OYE genes have been extensively characterized both biochemically and structurally (58,61,63).…”