Expression at 279 K, purification, crystallization and preliminary X-ray crystallographic analysis of a novel cold-active beta-1,4-D-mannanase from the Antarctic springtail Cryptopygus antarcticus

Abstract

The CaMan gene product from Cryptopygus antarcticus, which belongs to the glycoside hydrolase family 5 type beta-1,4-D-mannanases, has been crystallized using a precipitant solution consisting of 0.1 M Tris-HCl pH 8.5, 25%(w/v) polyethylene glycol 3350 by the microbatch crystallization method at 295 K. The CaMan protein crystal belonged to space group P2(1)2(1)2(1), with unit-cell parameters a = 73.40, b = 83.81, c = 163.55 angstrom. Assuming the presence of two molecules in the asymmetric unit, the solvent content was estimated to be about 61.29%. CaMan-mannopentaose (M5) complex crystals that were isomorphous to the CaMan crystals were obtained using the same mother liquor containing 1 mM M5.