Crystallization of Chlorella deoxyuridine triphosphatase

Abstract

Deoxyuridine triphosphatase (dUTPase) is a ubiquitous enzyme that has been widely studied owing to its function and evolutionary significance. The gene coding for the dUTPase from the Chlorella alga was codon-optimized and synthesized. The synthetic gene was expressed in Escherichia coli and recombinant core Chlorella dUTPase (chdUTPase) was purified. Crystallization of chdUTPase was performed by the repetitive hanging-drop vapor-diffusion method at 298 K with ammonium sulfate as the precipitant. In the presence of 2′-deoxyuridine-5′-[(α,β)-imido]triphosphate and magnesium, the enzyme produced die-shaped hexagonal R3 crystals with unit-cell parameters a = b = 66.9, c = 93.6 Å, γ = 120°. X-ray diffraction data for chdUTPase were collected to 1.6 Å resolution. The crystallization of chdUTPase with manganese resulted in very fragile clusters of needles.