Crystallization and preliminary characterization of dihydropteridine reductase from Dictyostelium discoideum

Abstract

Dihydropteridine reductase from Dictyostelium discoideum (dicDHPR) can produce D-threo-BH4 [6R-(1'R,2'R)-5,6,7,8-tetrahydrobiopterin], a stereoisomer of L-erythro-BH4, in the last step of tetrahydrobiopterin (BH4) recycling. In this reaction, DHPR uses NADH as a cofactor to reduce quinonoid dihydrobiopterin back to BH4. To date, the enzyme has been purified to homogeneity from many sources. In this report, the dicDHPR-NAD complex has been crystallized using the hanging-drop vapour-diffusion method with PEG 3350 as a precipitant. Rectangular-shaped crystals were obtained. Crystals grew to maximum dimensions of 0.4 x 0.6 x 0.1 mm. The crystal belonged to space group P2(1), with unit-cell parameters a = 49.81, b = 129.90, c = 78.76 angstrom, beta = 100.00 degrees, and contained four molecules in the asymmetric unit, forming two closely interacting dicDHPR-NAD dimers. Diffraction data were collected to 2.16 angstrom resolution using synchrotron radiation. The crystal structure has been determined using the molecular-replacement method.