Purification of a D-hydantoinase using a laboratory-scale Streamline phenyl column as the initial step

A D-hydantoinase from Thermus sp. was overexpressed in Escherichia coli and purified to homogeneity for subsequent crystallization. The purification w as performed with hydrophobic interaction chromatography as the capture ste p followed by anion-exchange chromatography and gel permeation chromatograp hy as intermediate purification and polishing steps, respectively. The hydr ophobic interaction step was done in fluidized bed mode in a laboratory-sca le Streamline column made from conventional laboratory equipment. The whole purification protocol could be finished within one day. The purified enzym e crystallizes. The crystals are suitable for X-ray protein structure analy sis and diffract to at least 2.3 Angstrom resolution. Complete data sets ha ve been measured up to 2.6 Angstrom resolution. The X-ray structure is curr ently being solved. (C) 2000 Published by Elsevier Science BN. All rights r eserved.