Cloning, expression, purification, crystallization and preliminary diffraction analysis of the C-terminal catalytic domain of human poly(ADP-ribose) polymerase

Abstract

Two fragments of the C-terminal catalytic domain of human poly(ADP-ribose) polymerase (catPARP), Met-catPARP and Gly-Ser-catPARP, were puri®ed and crystallized. Both catPARP crystals belong to space group C2, with almost the same unit-cell parameters. However, the shapes and harvest periods of both crystals were quite different owing to the slight mutation at the N-terminal position. Gly-Ser-catPARP was found to be more suitable for X-ray crystallography and crystals showed diffraction to at least 3.5 A Ê resolution.

Discovery of quinazolinone and quinoxaline derivatives as potent and selective poly(ADP‐ribose) polymerase‐1/2 inhibitors

“…The C-terminal catalytic domain of PARP (catPARP) from human was purified as described previously [12]. Crystals of catPARP complexed with 10 were obtained by the sitting drop vapor-diffusion method, adding equal volumes of concentrated protein solution (20 mg/ml) and a crystallization buffer (2.2-2.3 M (NH 4 ) 2 SO 4 , 1% v/v PEG400 and 100 mM Tris-HCl, pH 8.0) at 3:1 molar ratio of 10 to protein.…”