title: Channel formation by CarO, the carbapenem resistance-associated outer membrane protein of Acinetobacter baumannii.
abstract: It has been recently shown that resistance to both imipenem and meropenem in multidrug-resistant clinical strains of Acinetobacter baumannii is associated with the loss of a heat-modifiable 25/29-kDa outer membrane protein, called CarO. This study aimed to investigate the channel-forming properties of CarO. Mass spectrometry analyses of this protein band detected another 25-kDa protein (called Omp25), together with CarO. Both proteins presented similar physicochemical parameters (M(w) and pI). We overproduced and purified the two polypeptides as His-tagged recombinant proteins. Circular dichroism analyses demonstrated that the secondary structure of these proteins was mainly a beta-strand conformation with spectra typical of porins. We studied the channel-forming properties of proteins by reconstitution into artificial lipid bilayers. In these conditions, CarO induced ion channels with a conductance value of 110 pS in 1 M KCl, whereas the Omp25 protein did not form any channels, despite its suggested porin function. The pores formed by CarO showed a slight cationic selectivity and no voltage closure. No specific imipenem binding site was found in CarO, and this protein would rather form unspecific monomeric channels.
authors.affiliation: UMR 6522, CNRS, PBM, Plate-forme Protéomique IFRMP 23, Faculté des Sciences, Université de Rouen, F76821 Mont Saint Aignan Cedex, France.
authors.forename: Axel
authors.initials: A
authors.lastname: Siroy
authors.forename: Virginie
authors.initials: V
authors.lastname: Molle
authors.forename: Christelle
authors.initials: C
authors.lastname: Lemaître-Guillier
authors.forename: David
authors.initials: D
authors.lastname: Vallenet
authors.forename: Martine
authors.initials: M
authors.lastname: Pestel-Caron
authors.forename: Alain J
authors.initials: AJ
authors.lastname: Cozzone
authors.forename: Thierry
authors.initials: T
authors.lastname: Jouenne
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Dé
mesh_terms: D040981:Acinetobacter baumannii / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015780:Carbapenems / Q000378:metabolism / Q000494:pharmacology*; D024901:Drug Resistance, Multiple, Bacterial; D016680:Genome, Bacterial; D007473:Ion Channels / Q000737:chemistry*; D008051:Lipid Bilayers / Q000737:chemistry; D008826:Microbial Sensitivity Tests; D018272:Porins / Q000737:chemistry / Q000302:isolation & purification
chemical_list: D001425:Bacterial Outer Membrane Proteins; D015780:Carbapenems; D007473:Ion Channels; D008051:Lipid Bilayers; D018272:Porins