title: Combining metagenomics with metaproteomics and stable isotope probing reveals metabolic pathways used by a naturally occurring marine methylotroph.
abstract: A variety of culture-independent techniques have been developed that can be used in conjunction with culture-dependent physiological and metabolic studies of key microbial organisms in order to better understand how the activity of natural populations influences and regulates all major biogeochemical cycles. In this study, we combined deoxyribonucleic acid-stable isotope probing (DNA-SIP) with metagenomics and metaproteomics to characterize an uncultivated marine methylotroph that actively incorporated carbon from (13) C-labeled methanol into biomass. By metagenomic sequencing of the heavy DNA, we retrieved virtually the whole genome of this bacterium and determined its metabolic potential. Through protein-stable isotope probing, the RuMP cycle was established as the main carbon assimilation pathway, and the classical methanol dehydrogenase-encoding gene mxaF, as well as three out of four identified xoxF homologues were found to be expressed. This proof-of-concept study is the first in which the culture-independent techniques of DNA-SIP and protein-SIP have been used to characterize the metabolism of a naturally occurring Methylophaga-like bacterium in the marine environment (i.e. Methylophaga thiooxydans L4) and thus provides a powerful approach to access the genome and proteome of uncultivated microbes involved in key processes in the environment.
authors.affiliation: School of Environmental Sciences, University of East Anglia, Norwich Research Park, Norwich, NR4 7TJ, UK.
authors.forename: Carolina
authors.initials: C
authors.lastname: Grob
authors.affiliation: School of Environmental Sciences, University of East Anglia, Norwich Research Park, Norwich, NR4 7TJ, UK.
authors.forename: Martin
authors.initials: M
authors.lastname: Taubert
authors.affiliation: School of Environmental Sciences, University of East Anglia, Norwich Research Park, Norwich, NR4 7TJ, UK.
authors.forename: Alexandra M
authors.initials: AM
authors.lastname: Howat
authors.affiliation: School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich, NR4 7TJ, UK.
authors.forename: Oliver J
authors.initials: OJ
authors.lastname: Burns
authors.affiliation: Plymouth Marine Laboratory, Prospect Place, West Hoe, Plymouth, Devon, PL1 3DH, UK.
authors.forename: Joanna L
authors.initials: JL
authors.lastname: Dixon
authors.affiliation: Department of Isotope Biogeochemistry, Helmholtz-Centre for Environmental Research - UFZ, Leipzig, Germany.
authors.forename: Hans H
authors.initials: HH
authors.lastname: Richnow
authors.affiliation: Department of Proteomics, Helmholtz-Centre for Environmental Research-UFZ, Leipzig, Germany.
authors.forename: Nico
authors.initials: N
authors.lastname: Jehmlich
authors.affiliation: Department of Proteomics, Helmholtz-Centre for Environmental Research-UFZ, Leipzig, Germany.|Department of Metabolomics, Helmholtz-Centre for Environmental Research - UFZ, Leipzig, Germany.|Department of Chemistry and Bioscience, University of Aalborg, Fredrik Bajers Vej 7H, Aalborg East, 9220, Denmark.
authors.forename: Martin
authors.initials: M
authors.lastname: von Bergen
authors.affiliation: School of Life Sciences, University of Warwick, Coventry, CV4 7AL, UK.
authors.forename: Yin
authors.initials: Y
authors.lastname: Chen
authors.affiliation: School of Environmental Sciences, University of East Anglia, Norwich Research Park, Norwich, NR4 7TJ, UK.
authors.forename: J Colin
authors.initials: JC
authors.lastname: Murrell
mesh_terms: D000429:Alcohol Oxidoreductases / Q000235:genetics; D001483:Base Sequence; D018533:Biomass; D002244:Carbon / Q000378:metabolism; D004269:DNA, Bacterial / Q000235:genetics; D016680:Genome, Bacterial / Q000235:genetics; D007553:Isotope Labeling; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D056186:Metagenomics / Q000379:methods; D000432:Methanol / Q000378:metabolism*; D008969:Molecular Sequence Data; D044147:Piscirickettsiaceae / Q000235:genetics / Q000378:metabolism*; D020543:Proteome / Q000235:genetics; D040901:Proteomics / Q000379:methods; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012623:Seawater / Q000382:microbiology*; D017422:Sequence Analysis, DNA
chemical_list: D004269:DNA, Bacterial; D020543:Proteome; D012336:RNA, Ribosomal, 16S; D002244:Carbon; D000429:Alcohol Oxidoreductases; C019641:alcohol dehydrogenase (acceptor); D000432:Methanol