Biosynthesis of a 3,6-dideoxyhexose: crystallization and X-ray diffraction of CDP-6-deoxy-L-threo-D-­glycero-4-hexulose-3-dehydrase (E₁) for ascarylose biosynthesis

CDP-6-deoxy-L-threo-D-glycero-4-hexulose-3-dehydrase (E₁), along with its reductase (E₃), catalyzes the unusual C-3 deoxygenation of CDP-6-deoxy-L-­threo-D-glycero-4-hexulose to form CDP-3,6-dideoxy-L-threo-D-glycero-4-­hexulose in CDP-ascarylose biosynthesis [Chen et al. (1996), Biochemistry, 35, 16412–16420]. This dimeric [2Fe–2S] protein, cloned from the bacteria Yersinia pseudotuberculosis, is currently the only known example of an enzyme that uses a vitamin B₆-derived pyridoxamine 5′-phosphate (PMP) cofactor to carry out one-electron chemistry [Agnihotri & Liu (2001), Bioorg. Chem. 29, 234–257]. It also exhibits a [2Fe–2S] cluster-binding motif (C-X₅₇-C-X₁-C-X₇-C) which has not been observed previously [Agnihotri et al. (2004), Biochemistry, 43, 14265–14274] The recombinant 97.7 kDa dimer was crystallized in the trigonal space group P3₂, with unit-cell parameters a = b = 97.37, c = 142.2 Å, α = β = 90, γ = 120°. A data set has been collected to 1.9 Å resolution. A full MAD data set was also collected at the iron absorption edge that diffracted to 2.0 Å.