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.

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    Abstract

    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
    production.
    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
    URI: https://ueaeprints.uea.ac.uk/id/eprint/42417
    DOI:

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