Genetic analysis of DMSP metabolism in the marine Roseobacter clade

Kirkwood, Mark (2012) Genetic analysis of DMSP metabolism in the marine Roseobacter clade. Doctoral thesis, University of East Anglia.

Download (14MB) | Preview


    Genetic, biochemical, bioinformatic and molecular approaches were used to analyse
    microbial catabolism of dimethylsulfoniopropionate (DMSP), an abundant anti-stress
    compound made by marine phytoplankton.
    Members of the Roseobacter clade of marine α-proteobacteria may catabolise DMSP by two
    different routes; demethylation to form methylmercaptopropionate (MMPA), and cleavage by
    DMSP-lyases, yielding volatile dimethylsulfide (DMS) plus acrylate.
    The DMSP-lyase, DddP, was purified from Roseovarius nubinhibens ISM and characterised
    in vitro. Nuclear magnetic resonance spectroscopy and gas chromatography confirmed bona
    fide DMSP lyase activity and mutation of predicted active-site residues abolished DMS
    DddP was also detected in the fungal coral pathogen Aspergillus sydowii, likely acquired
    from bacteria by inter-Domain horizontal-gene-transfer.
    A new DMSP-lyase, DddW, was identified in another Roseobacter species, Ruegeria
    pomeroyi DSS-3, initially by microarray-based demonstrations that transcription of dddW
    was induced in cells grown with DMSP. An adjacent gene encoded the cognate
    transcriptional regulator. Escherichia coli cells that over-expressed DddW cleaved DMSP
    into DMS plus acrylate. Thus, Ruegeria pomeroyi has three DMSP-lyases, with DddP and
    DddQ being known already; mutational analyses showed that all three contributed to its
    DMSP-dependent DMS (Ddd+) phenotype.
    Moran’s laboratory had shown that the DMSP demethylase was encoded by R. pomeroyi
    dmdA. I unveiled intimate links between the demethylation and the cleavage pathway(s). A
    key player is acuI, which is co-transcribed with dmdA, both genes being induced by DMSP
    and, more markedly, the DMSP-catabolite, acrylate. Furthermore, AcuI- mutants failed to
    grow on acrylate as sole carbon source and were more sensitive to its toxic effects. AcuI-
    mutants failed to grow on DMSP so, surprisingly, Ruegeria likely uses lyase pathway(s) to
    grow on this compound. A potential regulatory gene, transcribed divergently from dmdA, was
    also identified.
    The microarray also, wholly unexpectedly, revealed a suite of cox genes involved in carbon
    monoxide oxidation that was up-regulated in response to DMS.

    Item Type: Thesis (Doctoral)
    Faculty \ School: Faculty of Science > School of Biological Sciences
    Depositing User: Mia Reeves
    Date Deposited: 16 May 2013 13:11
    Last Modified: 16 May 2013 13:11

    Actions (login required)

    View Item