The purified colicin S8 is a multimeric protein.
Bacteriocins have been isolated both as simple proteins and as proteins in association with carbohydrates, lipids, etc. Colicins are commonly inducible and extracellular. Their molecular masses range from 30 to 90 kDa. Pure colicin S8 was obtained in three steps from supernatant of induced cells: (i) Ammonium sulfate precipitation; (ii) anion exchange chromatography; and (iii) phenyl-Sepharose hydrophobic chromatography, either by preparative or fast performance liquid chromatography (FPLC) analytical purification procedure. In our hands, purified colicin S8 was an aggregation of extremely related polypeptides. Composition of those active fractions was the same: five polypeptides of molecular weight around 55 kDa. Behavior on molecular filtration indicated a molecular weight higher than 200 kDa. Similar results were obtained when purification was carried out through FPLC. Producing strains contain a single plasmid that encodes colicin S8; in minicells, this plasmid was shown to specify a 60 kDa polypeptide. We conclude that more than one form of colicin S8 exists. The forms are structurally related and can be recognized by antibodies raised against one of the polypeptides. Consistent with this conclusion, comparison of peptides produced after hydrolysis with chlorosuccinamide indicated that the active proteins contained both shared and unique components.
Concepción Curbelo
J L
JL
Department of Biology, Faculty of Sciences, University of the Andes, La Hechicera, Mérida, Venezuela.
Garcia Diaz
M E
ME
D000577:Amides / Q000737:chemistry / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry; D002845:Chromatography; D003087:Colicins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D004926:Escherichia coli / Q000378:metabolism*; D006868:Hydrolysis; D015151:Immunoblotting; D046911:Macromolecular Substances; D010957:Plasmids; D013386:Succinates
D000577:Amides; D001426:Bacterial Proteins; D003087:Colicins; D046911:Macromolecular Substances; D013386:Succinates; C011116:succinamide