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Mu-Like Prophage Strong Gyrase Site Sequences: Analysis of Properties Required for Promoting Efficient Mu DNA Replication. Mark Oram, 2004.The bacteriophage Mu genome contains a centrally located strong gyrase site (SGS) that is required for efficient prophage replication . To aid in studying the unusual properties of the SGS, we sought other gyrase sites that might be able to substitute for the SGS in Mu replication . Five candidate sites were obtained by PCR from Mu-like prophage sequences present in Escherichia coli O157:H7 Sakai, Haemophilus influenzae Rd, Salmonella enterica serovar Typhi CT18, and two strains of Neisseria meningitidis . Each of the sites was used to replace the natural Mu SGS to form recombinant prophages, and the effects on Mu replication and host lysis were determined . The site from the E . coli prophage supported markedly enhanced replication and host lysis over that observed with a Mu derivative lacking the SGS, those from the N . meningitidis prophages allowed a small enhancement, and the sites from the Haemophilus and Salmonella prophages gave none . Each of the candidate sites was cleaved specifically by E . coli DNA gyrase both in vitro and in vivo . Supercoiling assays performed in vitro, with the five sites or the Mu SGS individually cloned into a pUC19 reporter plasmid, showed that the Mu SGS and the E . coli or N . meningitidis sequences allowed an enhancement of processive, gyrase-dependent supercoiling, whereas the H . influenzae or Salmonella serovar Typhi sequences did not . While consistent with a requirement for enhanced processivity of supercoiling for a site to function in Mu replication, these data suggest that other factors are also important . The relevance of these observations to an understanding of the function of the SGS is discussed . Vibrio parahaemolyticus scrABC, a Novel Operon Affecting Swarming and Capsular Polysaccharide Regulation. Blaise R. Boles, 2002.Swarming is an adaptation of many bacteria to growth on surfaces . A search for genes controlling swarmer cell differentiation of Vibrio parahaemolyticus identified a novel three-gene operon that potentially encodes a pyridoxal-phosphate-dependent enzyme, an extracellular solute-binding protein, and a membrane-bound GGDEF- and EAL-motif sensory protein . The functions of these motifs, which are named after conserved amino acid sequences, are unknown, although the domains are found singly and in combination in a variety of bacterial signaling proteins . Studies with translational fusions supported the predicted localization of the gene products . When the operon was overexpressed, swarmer cell gene transcription was induced in liquid culture . Mutants with defects in any of the three genes exhibited decreased swarming and lateral flagellar (laf) gene expression . Complementation studies confirmed an operon organization and suggested that all three genes participated in laf regulation . The lesions that decreased swarming increased capsular polysaccharide (CPS) production, and overexpression of the operon inhibited transcription of the CPS gene cpsA . Thus, the scrABC locus appears to inversely regulate two gene systems that are pertinent to colonization of surface swarming and CPS .
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