The structure of the complex was refined to an R factor of 21.4% and R free ¼ 23.6% (for crystallographic statistics, see Supplementary Table S1). The final model includes protein residues 25–49 and 53–148, a total of 38 RNA nucleotides, 70 water molecules and 2 sulphate ions. N-terminal residues H25 and M26 are from the vector construct. Residues H25, D54, E80, F105, Q131 are modelled as partial side chains, whereas side chains of residues T111, S124, L133 and Q142 exhibit double conformations. Figures were prepared with PyMOL (http://pymol.sourceforge.net/) and GRASP 23 , and the RNA duplex curvature was estimated using CURVES 24 . Electrophoretic mobility shift assays The RNA duplexes (sequences available on request) were annealed as described 6 , and 5 0 -end-labelled with 32 P. The protein–RNA binding reactions contained 100fmol RNA duplex, 10 nmol (as monomer) full-length p19 and 5 ml 0.1 M KCl, 25 mM HEPES, 10 mM DTT, pH 7.6. 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The definition of generalized helicoidal parameters and of axis curvature for irregular nucleic acids. J. Biomol. Struct. Dyn. 6, 63–91 (1988). Supplementary Information accompanies the paper on www.nature.com/nature. Acknowledgements We thank members of the Patel laboratory for stimulating discussions and A. Teplov for assistance with data collection. This research was supported by NIH. We thank the personnel at beamline 14IDB of the Advanced Photon Source (APS) for their assistance. Use of this APS beamline was supported by the US Department of Energy, Basic Energy Sciences, Office of Science. Competing interests statement The authors declare that they have no competing financial interests. Correspondence and requests for materials should be addressed to D.P. (pateld@mskcc.org). Coordinates have been deposited in the Protein Data Bank under accession code 1R9F. .............................................................. A conspicuous nickel protein in microbial mats that oxidize methane anaerobically Martin Kru ¨ ger 1 , Anke Meyerdierks 1 , Frank Oliver Glo ¨ ckner 1 , Rudolf Amann 1 , Friedrich Widdel 1 , Michael Kube 2 , Richard Reinhardt 2 , Jo ¨ rg Kahnt 3 , Reinhard Bo ¨ cher 3 , Rudolf K. Thauer 3 & Seigo Shima 3 1 Max Planck Institute for Marine Microbiology, Celsiusstraße 1, 28359 Bremen, Germany 2 Max Planck Institute for Molecular Genetics, Ihnestraße, 14195 Berlin, Germany 3 Max Planck Institute for Terrestrial Microbiology, Karl-von-Frisch-Straße, 35043 Marburg, Germany ............................................................................................................................................................................. Anaerobic oxidation of methane (AOM) in marine sediments is an important microbial process in the global carbon cycle and in control of greenhouse gas emission. The responsible organisms supposedly reverse the reactions of methanogenesis 1–8 , but cul- tures providing biochemical proof of this have not been isolated. Here we searched for AOM-associated cell components in microbial mats from anoxic methane seeps in the Black Sea 9–11 . These mats catalyse AOM rather than carry out methanogenesis. We extracted a prominent nickel compound displaying the same absorption spectrum as the nickel cofactor F 430 of methyl-coenzyme M reductase, the terminal enzyme of methano- genesis 12 ; however, the nickel compound exhibited a higher molecular mass than F 430 . The apparent variant of F 430 was part of an abundant protein that was purified from the mat and that consists of three different subunits. Determined amino- terminal amino acid sequences matched a gene locus cloned from the mat. Sequence analyses revealed similarities to methyl- coenzyme M reductase from methanogenic archaea. The abun- dance of the nickel protein (7% of extracted proteins) in the mat suggests an important role in AOM. Large reservoirs of methane exceeding in mass conventional fossil fuels lie buried in deep, sulphate-depleted marine sediments 13,14 . Despite permanent geochemical or microbial production and upward migration, little of the methane ever escapes into the free ocean water. In the upper subsurface sediment, where sulphate but no oxygen is present, most methane is scavenged by anaerobic microbial oxidation 2,4,6,15–19 according to CH 4 þ SO 4 22 ! HCO 3 2 þ HS 2 þ H 2 O. Several in situ analyses based on isotope and lipid signatures 5,11,13,20–22 or phylogenetic marker DNA or RNA 5,7,10,11,14,22 have suggested that the microorganisms responsible for AOM represent special, hitherto uncultivated groups of archaea in association with sulphate-reducing bacteria. Most of these archaea belong to the ANME-1 and ANME-2 clusters. Both clusters are related to the Methanosarcinales 5,7,11,14,22 . The associated, tentative sulphate-reducing bacteria usually affiliate with the Desulfosarcina/ Desulfococcus cluster of the Deltaproteobacteria. Besides unique microorganisms, AOM involves intriguing mechanisms. There is still a dispute about how methane oxidation is linked to sulphate reduction 2,6,23,24 . Furthermore, the free energy gain from AOM (DG in situ estimated between 210 and 240 kJ mol 21 ) is one of the smallest known to fuel the metabolism of a microbial life-form 2,21,23,24 . Moreover, methane is chemically unreactive. Mechanisms for its biochemical activation are, there- fore, of particular interest. A natural system that seems to be well suited for a cultivation- independent biochemical study of AOM is microbial mats from the cold anoxic methane seeps of the northwestern Black Sea shelf 9–11 . These mats provide sufficient biomass to record a cytochrome spectrum 9 . They harbour archaea of the ANME-1 cluster that account for up to 70% of the cells detectable in situ 11 . Cells of the letters to nature NATURE | VOL 426 | 18/25 DECEMBER 2003 | www.nature.com/nature 878 © 2003 Nature Publishing Group