|
|
|
|
|
|
Characterization of the Upper Pathway Genes for Fluorene Metabolism in Terrabacter sp . Strain DBF63. Hiroshi Habe, 2004.Genes involved in the degradation of fluorene to phthalate were characterized in the fluorene degrader Terrabacter sp . strain DBF63 . The initial attack on both fluorene and 9-fluorenone was catalyzed by DbfA to yield 9-fluorenol and 1,1a-dihydroxy-1-hydro-9-fluorenone, respectively . The FlnB protein exhibited activities against both 9-fluorenol and 1,1a-dihydroxy-1-hydro-9-fluorenone to produce 9-fluorenone and 2'-carboxy-2,3-dihydroxybiphenyl, respectively . FlnD is a heteromeric protein encoded by flnD1 and ORF16, being a member of the class III two-subunit extradiol dioxygenase . FlnE was identified as a serine hydrolase for the meta-cleavage products that yield phthalate . Genomic Analysis of Clostridium perfringens Bacteriophage  3626, Which Integrates into guaA and Possibly Affects Sporulation.Markus Zimmer, 2002.Two temperate viruses, 3626 and
8533, have been isolated from lysogenic Clostridium perfringens strains . Phage
3626 was chosen for detailed analysis and was inspected by electron microscopy, protein profiling, and host range determination . For the first time, the nucleotide sequence of a bacteriophage infecting Clostridium species was determined . The virus belongs to the Siphoviridae family of the tailed phages, the order Caudovirales . Its genome consists of a linear double-stranded DNA molecule of 33,507 nucleotides, with invariable 3'-protruding cohesive ends of nine residues . Fifty open reading frames were identified, which are organized in three major life cycle-specific gene clusters . The genes required for lytic development show an opposite orientation and arrangement compared to the lysogeny control region . A function could be assigned to 19 gene products, based upon bioinformatic analyses, N-terminal amino acid sequencing, or experimental evidence . These include DNA-packaging proteins, structural components, a dual lysis system, a putative lysogeny switch, and proteins that are involved in replication, recombination, and modification of phage DNA . The presence of genes encoding a putative sigma factor related to sporulation-dependent sigma factors and a putative sporulation-dependent transcription regulator suggests a possible interaction of
3626 with onset of sporulation in C . perfringens . We found that the
3626 attachment site attP lies in a noncoding region immediately downstream of int . Integration of the viral genome occurs into the bacterial attachment site attB, which is located within the 3' end of a guaA homologue . This essential housekeeping gene is functionally independent of the integration status, due to reconstitution of its terminal codons by phage sequence .
Role of GlnK in NifL-Mediated Regulation of NifA Activity in Azotobacter vinelandii. Paul Rudnick, 2002.In several diazotrophic species of Proteobacteria, PII signal transduction proteins have been implicated in the regulation of nitrogen fixation in response to NH4+ by several mechanisms . In Azotobacter vinelandii, expression of nifA, encoding the nif-specific activator, is constitutive, and thus, regulation of NifA activity by the flavoprotein NifL appears to be the primary level of nitrogen control . In vitro and genetic evidence suggests that the nitrogen response involves the PII-like GlnK protein and GlnD (uridylyltransferase/uridylyl-removing enzyme), which reversibly uridylylates GlnK in response to nitrogen limitation . Here, the roles of GlnK and GlnK-UMP in A . vinelandii were studied to determine whether the Nif - phenotype of glnD strains was due to an inability to modify GlnK, an effort previously hampered because glnK is an essential gene in this organism . A glnKY51F mutation, encoding an unuridylylatable form of the protein, was stable only in a strain in which glutamine synthetase activity is not inhibited by NH4+, suggesting that GlnK-UMP is required to signal adenylyltransferase/adenylyl-removing enzyme-mediated deadenylylation . glnKY51F strains were significantly impaired for diazotrophic growth and expression of a nifH-lacZ fusion . NifL interacted with GlnK and GlnKY51F in a yeast two-hybrid system . Together, these data are consistent with those obtained from in vitro experiments (Little et al., EMBO J., 19:6041–6050, 2000) and support a model for regulation of NifA activity in which unmodified GlnK stimulates NifL inhibition and uridylylation of GlnK in response to nitrogen limitation prevents this function . This model is distinct from one proposed for the related bacterium Klebsiella pneumoniae, in which unmodified GlnK relieves NifL inhibition instead of stimulating it . Mutant Analysis of the Escherichia coli FhuA Protein Reveals Sites of FhuA Activity. Franziska Endriß, 2003.The FhuA outer membrane protein of Escherichia coli actively transports ferrichrome, albomycin, and rifamycin CGP 4832, and confers sensitivity to microcin J25, colicin M, and the phages T1, T5, and  80 . Guided by the FhuA crystal structure and derived predictions on how FhuA might function, mutants were isolated in the cork domain (residues 1 to 160) and in the ß-barrel domain (residues 161 to 714) . Deletion of the TonB box (residues 7 to 11) completely inactivated all TonB-dependent functions of FhuA . Fixation of the cork to turn 7 of the barrel through a disulfide bridge between introduced C27 and C533 residues abolished ferrichrome transport, which was restored by reduction of the disulfide bond . Deletion of residues 24 to 31, including the switch helix (residues 24 to 29), which upon binding of ferrichrome to FhuA undergoes a large structural transition (17 Å) and exposes the N terminus of FhuA (TonB box) to the periplasm, reduced FhuA transport activity (79% of the wild-type activity) but conferred full sensitivity to colicin M and the phages . Duplication of residues 23 to 30 or deletion of residues 13 to 20 resulted in FhuA derivatives with properties similar to those of FhuA with a deletion of residues 24 to 31 . However, a frameshift mutation that changed QSEA at positions 18 to 21 to KKAP abolished almost completely most of FhuA's activities . The conserved residues R93 and R133 among energy-coupled outer membrane transporters are thought to fix the cork to the ß-barrel by forming salt bridges to the conserved residues E522 and E571 of the ß-barrel . Proteins with the E522R and E571R mutations were inactive, but inactivity was not caused by repulsion of R93 by R522 and R571 and of R133 by R571 . Point mutations in the cork at sites that move or do not move upon the binding of ferrichrome had no effect or conferred only slightly reduced activities . It is concluded that the TonB box is essential for FhuA activity . The TonB box region has to be flexible, but its distance from the cork domain can greatly vary . The removal of salt bridges between the cork and the barrel affects the structure but not the function of FhuA .
Disruption of the CAR1 Gene Encoding Arginase Enhances Freeze Tolerance of the Commercial Baker's Yeast Saccharomyces cerevisiae. Jun Shima, 2003.The effect of intracellular charged amino acids on freeze tolerance in doughs was determined by constructing homozygous diploid arginase-deficient mutants of commercial baker's yeast . An arginase mutant accumulated higher levels of arginine and/or glutamate and showed increased leavening ability during the frozen-dough baking process, suggesting that disruption of the CAR1 gene enhances freeze tolerance . Anaerobic Ammonium Oxidation Measured in Sediments along the Thames Estuary, United Kingdom. Mark Trimmer, 2003.Until recently, denitrification was thought to be the only significant pathway for N2 formation and, in turn, the removal of nitrogen in aquatic sediments . The discovery of anaerobic ammonium oxidation in the laboratory suggested that alternative metabolisms might be present in the environment . By using a combination of 15N-labeled NH4+, NO3-, and NO2- (and 14N analogues), production of 29N2 and 30N2 was measured in anaerobic sediment slurries from six sites along the Thames estuary . The production of 29N2 in the presence of 15NH4+ and either 14NO3- or 14NO2- confirmed the presence of anaerobic ammonium oxidation, with the stoichiometry of the reaction indicating that the oxidation was coupled to the reduction of NO2- . Anaerobic ammonium oxidation proceeded at equal rates via either the direct reduction of NO2- or indirect reduction, following the initial reduction of NO3- . Whether NO2- was directly present at 800 µM or it accumulated at 3 to 20 µM (from the reduction of NO3-), the rate of 29N2 formation was not affected, which suggested that anaerobic ammonium oxidation was saturated at low concentrations of NO2- . We observed a shift in the significance of anaerobic ammonium oxidation to N2 formation relative to denitrification, from 8% near the head of the estuary to less than 1% at the coast . The relative importance of anaerobic ammonium oxidation was positively correlated (P < 0.05) with sediment organic content . This report of anaerobic ammonium oxidation in organically enriched estuarine sediments, though in contrast to a recent report on continental shelf sediments, confirms the presence of this novel metabolism in another aquatic sediment system .
|
© 2005
Transgalactic Ltd (manufacturer of Bioscreen C software) |
Privacy Statement | P.O. Box
1393, 00101 Helsinki, Finland,
Last modified: May 25, 2005
| ||||||
