Crystallization and preliminary X‐ray analysis of 5‐keto‐d‐gluconate reductase from Gluconobacter suboxydans IFO12528 complexed with 5‐keto‐d‐gluconate and NADPH
NADPH‐dependent 5‐keto‐d‐gluconate reductase from Gluconobacter suboxydans IFO12528 (5KGR) catalyzes oxidoreduction between 5‐keto‐d‐gluconate and d‐gluconate with high specificity. 5KGR was expressed in Escherichia coli, purified and crystallized with 5‐keto‐d‐gluconate and NADPH using the sitting‐drop vapour‐diffusion method at 288 K. A crystal of the 5KGR–NADPH complex was obtained using reservoir solution containing PEG 4000 as a precipitant and diffracted X‐rays to 1.75 Å resolution. The crystal of the complex belonged to space group P42212, with unit‐cell parameters a = b = 128.6, c = 62.9 Å. A crystal of the 5KGR–NADPH–5‐keto‐d‐gluconate complex was prepared by soaking the 5KGR–NADPH complex crystal in reservoir solution supplemented with 100 mM 5‐keto‐d‐gluconate and 10 mM NADPH for 20 min and diffracted X‐rays to 2.26 Å resolution. The crystal of the ternary complex belonged to space group P42212, with unit‐cell parameters a = b = 128.7, c = 62.5 Å. Both crystals contained two molecules in the asymmetric unit.
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