Towards structural studies of the old yellow enzyme homologue SYE4 from Shewanella oneidensis and its complexes at atomic resolution

• Published in: Acta crystallographica. Section F, Structural biology and crystallization communications Structural biology and crystallization communications, 2010-01, Vol.66 (1), p.85-90
• Main Authors: Elegheert, Jonathan, Van Den Hemel, Debbie, Dix, Ina, Stout, Jan, Van Beeumen, Jozef, Brigé, Ann, Savvides, Savvas N.
• Format: Article
• Language: English
• Subjects: Cloning, Molecular; Crystallization; Crystallization Communications; Crystallography, X-Ray; Escherichia coli; NADPH Dehydrogenase - chemistry; old yellow enzyme homologues; Shewanella - enzymology; Shewanella oneidensis; SYE4
• ISSN: 1744-3091; 1744-3091; 2053-230X
• DOI: 10.1107/S1744309109050386

Shewanella oneidensis is an environmentally versatile Gram‐negative γ‐proteobacterium that is endowed with an unusually large proteome of redox proteins. Of the four old yellow enzyme (OYE) homologues found in S. oneidensis, SYE4 is the homologue most implicated in resistance to oxidative stress. SYE4 was recombinantly expressed in Escherichia coli, purified and crystallized using the hanging‐drop vapour‐diffusion method. The crystals belonged to the orthorhombic space group P212121 and were moderately pseudo‐merohedrally twinned, emulating a P422 metric symmetry. The native crystals of SYE4 were of exceptional diffraction quality and provided complete data to 1.10 Å resolution using synchrotron radiation, while crystals of the reduced enzyme and of the enzyme in complex with a wide range of ligands typically led to high‐quality complete data sets to 1.30–1.60 Å resolution, thus providing a rare opportunity to dissect the structure–function relationships of a good‐sized enzyme (40 kDa) at true atomic resolution. Here, the attainment of a number of experimental milestones in the crystallographic studies of SYE4 and its complexes are reported, including isolation of the elusive hydride–Meisenheimer complex.