Purification and characterization of colicin D.
Colicin D-CA23, obtained by sonic treatment of mitomycin C-induced cells of Escherichia coli K-12 W1485 (colD), was purified by ammonium sulfate precipitation, gel filtration on Sephadex G200, ion-exchange chromatography on diethylaminoethyl cellulose, and isoelectrofocusing. Polyacrylamide-gel electrophoresis, sedimentation velocity analysis, and antigenic analysis indicated that the preparation was homogeneous. Colicin D is composed entirely of amino acids and hence is a simple protein uncomplexed with lipid or lipopolysaccharide. It contains six residues of cysteine per molecule. The molecular weight of colicin D is approximately 92,000, as determined by sodium dodecyl sulfate-polyacrylamide-gel electrophoresis and gel filtration on Sephadex G200. Its sedimentation coefficient is 4.41S. The behavior of colicin D in solutions of sodium dodecyl sulfate and 2-mercaptoethanol indicates that it does not consist of subunits and exists as a single polypeptide chain. Its high molecular weight and presence of six cysteine residues per molecule distinguish colicin D from all colicins previously described. Although colicins D and E3 have similar modes of action, their gross molecular properties are entirely different.
D000596:Amino Acids / Q000032:analysis; D000645:Ammonium Sulfate; D000818:Animals; D000942:Antigens, Bacterial / Q000032:analysis; D002474:Cell-Free System; D011232:Chemical Precipitation; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D003087:Colicins / Q000032:analysis / Q000096:biosynthesis / Q000302:isolation & purification; D003227:Conjugation, Genetic; D003470:Culture Media; D004589:Electrophoresis, Disc; D004926:Escherichia coli / Q000254:growth & development / Q000276:immunology / Q000378:metabolism; D007106:Immune Sera; D005779:Immunodiffusion; D007525:Isoelectric Focusing; D008970:Molecular Weight; D009500:Neutralization Tests; D011817:Rabbits; D014461:Ultracentrifugation
D000596:Amino Acids; D000942:Antigens, Bacterial; D003087:Colicins; D003470:Culture Media; D007106:Immune Sera; D000645:Ammonium Sulfate