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Applied and Environmental Microbiology, September 2003, p. 5483-5491, Vol. 69, No. 9
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.9.5483-5491.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Identification of Methyl Coenzyme M Reductase A (mcrA) Genes Associated with Methane-Oxidizing Archaea

Steven J. Hallam,1 Peter R. Girguis,1 Christina M. Preston,1 Paul M. Richardson,2 and Edward F. DeLong1*

Monterey Bay Aquarium Research Institute, Moss Landing, California 95039-9644,1 The Joint Genome Institute, Walnut Creek, California 945982

Received 28 March 2003/ Accepted 25 June 2003

Phylogenetic and stable-isotope analyses implicated two methanogen-like archaeal groups, ANME-1 and ANME-2, as key participants in the process of anaerobic methane oxidation. Although nothing is known about anaerobic methane oxidation at the molecular level, the evolutionary relationship between methane-oxidizing archaea (MOA) and methanogenic archaea raises the possibility that MOA have co-opted key elements of the methanogenic pathway, reversing many of its steps to oxidize methane anaerobically. In order to explore this hypothesis, the existence and genomic conservation of methyl coenzyme M reductase (MCR), the enzyme catalyzing the terminal step in methanogenesis, was studied in ANME-1 and ANME-2 archaea isolated from various marine environments. Clone libraries targeting a conserved region of the alpha subunit of MCR (mcrA) were generated and compared from environmental samples, laboratory-incubated microcosms, and fosmid libraries. Four out of five novel mcrA types identified from these sources were associated with ANME-1 or ANME-2 group members. Assignment of mcrA types to specific phylogenetic groups was based on environmental clone recoveries, selective enrichment of specific MOA and mcrA types in a microcosm, phylogenetic congruence between mcrA and small-subunit rRNA tree topologies, and genomic context derived from fosmid sequences. Analysis of the ANME-1 and ANME-2 mcrA sequences suggested the potential for catalytic activity based on conservation of active-site amino acids. These results provide a basis for identifying methanotrophic archaea with mcrA sequences and define a functional genomic link between methanogenic and methanotrophic archaea.

* Corresponding author. Mailing address: Monterey Bay Aquarium Research Institute, 7700 Sandholdt Rd., Moss Landing, CA 95039. Phone: (831) 775-1843. Fax: (831) 775-1620. E-mail: delong{at}mbari.org.

Applied and Environmental Microbiology, September 2003, p. 5483-5491, Vol. 69, No. 9
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.9.5483-5491.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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