RESUMO
Cells of Flavobacterium johnsoniae move over surfaces by a process known as gliding motility. The mechanism of this form of motility is not known. Cells of F. johnsoniae propel latex spheres along their surfaces, which is thought to be a manifestation of the motility machinery. Three of the genes that are required for F. johnsoniae gliding motility, gldA, gldB, and ftsX, have recently been described. Tn4351 mutagenesis was used to identify another gene, gldD, that is needed for gliding. Tn4351-induced gldD mutants formed nonspreading colonies, and cells failed to glide. They also lacked the ability to propel latex spheres and were resistant to bacteriophages that infect wild-type cells. Introduction of wild-type gldD into the mutants restored motility, ability to propel latex spheres, and sensitivity to bacteriophage infection. gldD codes for a cytoplasmic membrane protein that does not exhibit strong sequence similarity to proteins of known function. gldE, which lies immediately upstream of gldD, encodes another cytoplasmic membrane protein that may be involved in gliding motility. Overexpression of gldE partially suppressed the motility defects of a gldB point mutant, suggesting that GldB and GldE may interact. GldE exhibits sequence similarity to Borrelia burgdorferi TlyC and Salmonella enterica serovar Typhimurium CorC.
Assuntos
Proteínas de Bactérias/genética , Flavobacterium/genética , Proteínas de Membrana/genética , Sequência de Aminoácidos , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/fisiologia , Bacteriófagos/patogenicidade , Sequência de Bases , Clonagem Molecular , Elementos de DNA Transponíveis , DNA Bacteriano , Flavobacterium/metabolismo , Flavobacterium/fisiologia , Flavobacterium/virologia , Expressão Gênica , Genes Bacterianos , Teste de Complementação Genética , Proteínas de Membrana/isolamento & purificação , Proteínas de Membrana/metabolismo , Proteínas de Membrana/fisiologia , Dados de Sequência Molecular , Mutagênese Insercional , RNA Bacteriano/análise , Análise de Sequência de DNA , Homologia de Sequência de AminoácidosRESUMO
Shewanella putrefaciens is a facultative anaerobe that can use metal oxides as terminal electron acceptors during anaerobic respiration. Two proteins, MtrB and Cct, have been identified that are specifically involved in metal reduction. Analysis of S. putrefaciens mutants deficient in metal reduction led to the identification of two additional proteins that are involved in this process. MtrA is a periplasmic decahaem c-type cytochrome that appears to be part of the electron transport chain, which leads to Fe(III) and Mn(IV) reduction. MtrC is an outer membrane decahaem c-type cytochrome that appears to be required for the activity of the terminal Fe(III) reductase. Membrane fractions of mutants deficient in MtrC exhibited a decreased level of Fe(III) reduction compared with the wild type. We suggest that MtrC may be a component of the terminal reductase or may be required for its assembly.
Assuntos
Proteínas da Membrana Bacteriana Externa/metabolismo , Grupo dos Citocromos c/metabolismo , FMN Redutase , Compostos Férricos/metabolismo , Shewanella putrefaciens/enzimologia , Anaerobiose , Proteínas da Membrana Bacteriana Externa/genética , Sequência de Bases , Western Blotting , Grupo dos Citocromos c/genética , Deleção de Genes , Manganês/metabolismo , Dados de Sequência Molecular , NADH NADPH Oxirredutases/metabolismo , Oxirredução , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Shewanella putrefaciens/genética , Shewanella putrefaciens/crescimento & desenvolvimentoRESUMO
The mechanism of bacterial gliding motility (active movement over surfaces without the aid of flagella) is not known. A large number of mutants of the gliding bacterium Flavobacterium johnsoniae (Cytophaga johnsonae) with defects in gliding motility have been previously isolated, and genetic techniques to analyze these mutants have recently been developed. We complemented a nongliding mutant of F. johnsoniae (UW102-99) with a library of wild-type DNA by using the shuttle cosmid pCP26. The complementing plasmid (pCP200) contained an insert of 26 kb and restored gliding motility to 4 of 50 independently isolated nongliding mutants. A 1.9-kb fragment which encompassed two genes, gldB and gldC, complemented all four mutants. An insertion mutation in gldB was polar on gldC, suggesting that the two genes form an operon. Disruption of the chromosomal copy of gldB in wild-type F. johnsoniae UW101 eliminated gliding motility. Introduction of the gldBC operon, or gldB alone, restored motility. gldB appears to be essential for F. johnsoniae gliding motility. It codes for a membrane protein that does not exhibit strong sequence similarity to other proteins in the databases. gldC is not absolutely required for gliding motility, but cells that do not produce GldC form colonies that spread less well than those of the wild type. GldC is a soluble protein and has weak sequence similarity to the fungal lectin AOL.
Assuntos
Flavobacterium/genética , Flavobacterium/fisiologia , Genes Bacterianos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Bacteriófagos/fisiologia , Fracionamento Celular , Clonagem Molecular , Cosmídeos/genética , Cytophaga/genética , Cytophaga/fisiologia , Elementos de DNA Transponíveis , Flavobacterium/virologia , Teste de Complementação Genética , Dados de Sequência Molecular , Movimento , Mutagênese Insercional , Óperon/genética , Mutação PuntualRESUMO
The mechanism of bacterial gliding motility (active movement over surfaces without the aid of flagella) is not known. A large number of nonmotile mutants of the gliding bacterium Flavobacterium johnsoniae (Cytophaga johnsonae) have been previously isolated, and genetic techniques to analyze these mutants have recently been developed. We complemented a nonmotile mutant of F. johnsoniae (UW102-09) with a library of wild-type DNA by using the shuttle cosmid pCP17. The complementing plasmid (pCP100) contained an insert of 13 kbp, and restored motility to 4 of 61 independently isolated nonmotile mutants. A 1.3-kbp fragment that encompassed a single ORF, gldA, complemented all four mutants. Disruption of the chromosomal copy of gldA in wild-type F. johnsoniae UW101 eliminated gliding motility. The predicted protein produced by gldA has strong sequence similarity to ATP binding cassette transport proteins.
