A PROCEDURE FOR RENATURATION AND PURIFICATION OF THE EXTRACELLULAR SERRATIA-MARCESCENS NUCLEASE FROM GENETICALLY-ENGINEERED ESCHERICHIA-COLI

Overproduction of the extracellular Serratia marcescens nuclease in Escherichia coli results in aggregation and sequestration of a large amount of the protein in inclusion bodies. Only a relatively small amount is secreted into the medium from which it can be purified following established procedures. The cell-associated insoluble protein can be solubilized in 6 M urea after breaking up the cells by sonication. Renaturation is achieved by dilution or dialysis. Subsequent phosphocellulose chromatography yields a homogeneous protein preparation which is shown by a variety of biochemical and biophysical analyses to be indistinguishable from conventionally prepared material. The high yield (>10 mg/500-ml culture) and the ease of preparation (2 to 3 days) make this an attractive alternative to previously described procedures. © 1994 Academic Press. All rights reserved.