Molecular characterization of the Serratia marcescens OmpF porin, and analysis of S. marcescens OmpF and OmpC osmoregulation.
Serratia marcescens is a nosocomial pathogen with a high incidence of beta-lactam resistance. Reduced amounts of outer-membrane porins have been correlated with increased resistance to beta-lactams but only one porin, OmpC, has been characterized at the molecular level. In this study we present the molecular characterization of a second porin, OmpF, and an analysis of the expression of S. marcescens porins in response to various environmental changes. Two porins were isolated from the outer membrane using urea-SDS-PAGE and the relative amounts were shown to be influenced by the osmolarity of the medium and the presence of salicylate. From a S. marcescens genomic DNA library an 8 kb EcoRI fragment was isolated that hybridized with an oligonucleotide encoding the published N-terminal amino acid sequence of the S. marcescens 41 kDa porin. A 41 kDa protein was detected in the outer membrane of Escherichia coli NM522 carrying the cloned S. marcescens DNA. The cloned gene was sequenced and shown to code for a protein that shared 60-70% identity with other known OmpF and OmpC sequences. The upstream DNA sequence of the S. marcescens gene was similar to the corresponding E. coli ompF sequence; however, a regulatory element important in repression of E. coli ompF at high osmolarity was absent. The cloned S. marcescens OmpF in E. coli increased in expression in conditions of high osmolarity. The potential involvement of micF in the observed osmoregulation of S. marcescens porins is discussed.
Hutsul
Jo-Anne
JA
Department of Microbiology, University of Manitoba, Winnipeg, Manitoba, Canada R3T 2N2.
Worobec
Elizabeth
E
Department of Microbiology, University of Manitoba, Winnipeg, Manitoba, Canada R3T 2N2.
D001483:Base Sequence; D003001:Cloning, Molecular; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D006801:Humans; D008969:Molecular Sequence Data; D018272:Porins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D015183:Restriction Mapping; D012459:Salicylates / Q000494:pharmacology; D020156:Salicylic Acid; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid; D016868:Serratia Infections / Q000382:microbiology; D012706:Serratia marcescens / Q000187:drug effects / Q000235:genetics*; D013045:Species Specificity; D014882:Water-Electrolyte Balance
D018272:Porins; D012459:Salicylates; D020156:Salicylic Acid