title: Protease activity of outer membrane protein OmpT in clinical E.coli isolates--studies using translation initiation factor IF2 as target protein.
abstract: During purification of the translation initiation factor IF2 from ompT+ strains of Escherichia coli the IF2 is partially degraded in the presence of membrane debris during the first steps of purification. This is a result of proteolytic cleavage by outer membrane protease OmpT [1]. Here we have investigated the activity of OmpT in 51 clinical E. coli isolates of human origin, by a time dependent OmpT activity assay using IF2 as target protein. The activity of OmpT in the outer cell membrane is highly variable among wild type E.coli strains, ranging from no detectable activity in 65% of the strains to a very high activity in 5 strains (10%). The OmpT activity is closely related to the assay temperature and to the growth temperature of the cells, and can be reduced or even eliminated by lowering the temperature of growth. The results open the possibility of using non-denaturing gel electrophoresis of crude cell lysates as a screening method in population genetic studies of initiation factor IF2 and other cytoplasmic proteins which are cleaved by OmpT.
authors.affiliation: Department of Chemistry, Aarhus University, Denmark.
authors.forename: S
authors.initials: S
authors.lastname: Steffensen
authors.forename: A B
authors.initials: AB
authors.lastname: Poulsen
authors.forename: K K
authors.initials: KK
authors.lastname: Mortensen
authors.forename: B
authors.initials: B
authors.lastname: Korsager
authors.forename: H U
authors.initials: HU
authors.lastname: Sperling-Petersen
mesh_terms: D002455:Cell Division; D002462:Cell Membrane / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology* / Q000254:growth & development; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism*; D006801:Humans; D012697:Serine Endopeptidases / Q000378:metabolism*; D013696:Temperature
chemical_list: D015852:Eukaryotic Initiation Factor-2; D012697:Serine Endopeptidases; C057840:omptin outer membrane protease