PURIFICATION OF VIRUSES AND MACROMOLECULAR ASSEMBLIES FOR STRUCTURAL INVESTIGATIONS USING A NOVEL ION-EXCHANGE METHOD

We describe a novel ion exchange chromatographic technique suitable fo r large-scale preparation of viruses and other biomacromolecular assem blies in highly purified form. The method, which utilizes anion exchan ge on either of two commercially available cellulose cartridges, is ap plied to the Escherichia coli bacteriophage PRD1. Viral particles elut ed from both QMA and DEAE cartridges retain infectivity and exhibit gr eater homogeneity of composition, as judged by gel electrophoresis and electron microscopy, than particles purified by rate zonal sucrose gr adient centrifugation. The ion exchange protocols are rapid, requiring less than 15 min elution time, and permit retrieval of the purified v iral particles at high concentration in aqueous media without centrifu gal pelleting. The present method is particularly well suited to the p reparation of milligram to decigram quantities of virus, sufficient fo r many biophysical structural analyses, including investigations by so lution spectroscopic and crystal diffraction techniques. The feasibili ty and advantages of the ion exchange chromatographic procedure are de monstrated by application of laser Raman spectroscopy to ion exchange purified PRD1 virions and subviral assemblies, (C) 1994 Academic Press , Inc.