title: A procedure for renaturation and purification of the extracellular Serratia marcescens nuclease from genetically engineered Escherichia coli.
abstract: 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.
authors.affiliation: Institut für Biochemie, Fachbereich Biologie, Justus-Liebig-Universität, Giessen, Germany.
authors.forename: P
authors.initials: P
authors.lastname: Friedhoff
authors.forename: O
authors.initials: O
authors.lastname: Gimadutdinow
authors.forename: T
authors.initials: T
authors.lastname: Rüter
authors.forename: W
authors.initials: W
authors.lastname: Wende
authors.forename: C
authors.initials: C
authors.lastname: Urbanke
authors.forename: H
authors.initials: H
authors.lastname: Thole
authors.forename: A
authors.initials: A
authors.lastname: Pingoud
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000302:isolation & purification*; D002482:Cellulose / Q000031:analogs & derivatives; D002846:Chromatography, Affinity; D004706:Endodeoxyribonucleases / Q000096:biosynthesis / Q000302:isolation & purification*; D004722:Endoribonucleases / Q000096:biosynthesis / Q000302:isolation & purification*; D004926:Escherichia coli; D011489:Protein Denaturation; D017510:Protein Folding; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification*; D012706:Serratia marcescens / Q000201:enzymology* / Q000235:genetics; D014508:Urea
chemical_list: D001426:Bacterial Proteins; D011993:Recombinant Fusion Proteins; D014508:Urea; D002482:Cellulose; C008881:phosphocellulose; D004706:Endodeoxyribonucleases; D004722:Endoribonucleases; C021835:Serratia marcescens nuclease