Dark matter in a deep-sea vent and in human mouth.

Summer used to be a slow time for genomics news. This year, even the summer heat failed to stem the influx of new completely sequenced microbial genomes. The latest list (Table 1) includes genomes from a number of environmental bacteria (Chen ; Nakagawa ), four methanogenic archaea, as well as the finished genome of the red alga Cyanidioschyzon merolae, the first 100% complete eukaryotic genome (Nozaki ). However, this time the most striking news comes from an unfinished genome, the genome sequence of the first representative of the enigmatic TM7 phylum (Hugenholtz ). So far, no member of this widespread phylum has been isolated in pure culture and the genome sequence, even an incomplete one, provides the first glimpse into the physiology of this biological ‘dark matter’ (Marcy ).

Table 1

Recently completed microbial genomes (June–August 2007).

Species name	Taxonomy	GenBank accession	Genome size, bp	Proteins (total)	Sequencing centrea	Reference
New organisms
Cyanidioschyzon merolae	Eukaryota, Rhodophyta	AP006483–AP006502	16 546 747 (total)	4775	Univ. Tokyo	Nozaki et al. (2007)
Methanococcus aeolicus	Euryarchaeota	CP000743	1 569 500	1490	JGI	Unpublished
Methanococcus vannielii	Euryarchaeota	CP000742	1 720 048	1678	JGI	Unpublished
Candidatus Methanoregula boonei	Euryarchaeota	CP000780	2 542 943	2450	JGI	Unpublished
Kineococcus radiotolerans	Actinobacteria	CP000750	4 761 183	4497	JGI	Unpublished
		CP000752	12 917
Bacteroides vulgatus	Bacteroidetes	CP000139	5 163 189	4065	WashU	Xu et al. (2007)
Flavobacterium psychrophilum	Bacteroidetes	AM398681	2 861 988	2412	INRA – Jouy-en-Josas	Duchaud et al. (2007)
Parabacteroides distasonis	Bacteroidetes	CP000140	4 811 379	3850	WashU	Xu et al. (2007)
Alkaliphilus metalliredigens	Firmicutes	CP000724	4 929 566	4625	JGI	Unpublished
Bacillus amyloliquefaciens	Firmicutes	CP000560	3 918 589	3693	U. Göttingen	Chen et al. (2007)
Clostridium beijerinckii	Firmicutes	CP000721	6 000 632	5020	JGI	Unpublished
Clostridium kluyveri	Firmicutes	CP000673	3 964 618	3913	U. Göttingen	Unpublished
		CP000674	59 182
Ochrobactrum anthropi	α-Proteobacteria	CP000758–CP000763	5 205 777 (total)	4799	JGI	Unpublished
Parvibaculum lavamentivorans	α-Proteobacteria	CP000774	3 914 745	3636	JGI	Unpublished
Sinorhizobium medicae	α-Proteobacteria	CP000738–CP000741	6 817 576	6213	JGI	Unpublished
Xanthobacter autotrophicus	α-Proteobacteria	CP000781	5 308 934	5035	JGI	Unpublished
			316 164
Janthinobacterium sp. Marseille	β-Proteobacteria	CP000269	4 110 251	3697	CNRS-Marseille	Audic et al. (2007)
Actinobacillus succinogenes	γ-Proteobacteria	CP000746	2 319 663	2079	JGI	Unpublished
Klebsiella pneumoniae	γ-Proteobacteria	CP000647	5 694 894	5187	WashU	Unpublished
Marinomonas sp. MWYL1	γ-Proteobacteria	CP000749	5 100 344	4439	JGI	Unpublished
Anaeromyxobacter sp. Fw109-5	δ-Proteobacteria	CP000769	5 277 990	4466	JGI	Unpublished
Campylobacter curvus	ε-Proteobacteria	CP000767	1 971 264	1931	JCVI	Unpublished
Campylobacter hominis	ε-Proteobacteria	CP000776	1 711 273	1682	JCVI	Unpublished
Nitratiruptor sp. SB155-2	ε-Proteobacteria	AP009179	1 877 931	1857	JAMSTEC	Nakagawa et al. (2007)
Sulfurovum sp. NBC37-1	ε-Proteobacteria	AP009179	2 562 277	2466	JAMSTEC	Nakagawa et al. (2007)
Fervidobacterium nodosum	Thermotogae	CP000771	1 948 941	1750	JGI	Unpublished
Thermosipho melanesiensis	Thermotogae	CP000716	1 915 238	1879	JGI	Unpublished
New strains
Methanococcus maripaludis C7	Euryarchaeota	CP000745	1 772 694	1788	JGI	Unpublished
Mycobacterium tuberculosis F11	Actinobacteria	CP000717	4 424 435	3941	Broad Institute	Unpublished
Bacillus cereus ssp. cytotoxis NVH 391-98	Firmicutes	CP000764	4 087 024	3844	JGI	Lapidus et al. (2007)
		CP000765	7 135
Clostridium botulinum A str. ATCC 19397	Firmicutes	CP000726	3 863 450	3552	Los Alamos	Unpublished
Clostridium botulinum A str. Hall	Firmicutes	CP000727	3 760 560	3407	Los Alamos	Unpublished
Clostridium botulinum F str. Langeland	Firmicutes	CP000728	3 995 387	3659	Los Alamos	Unpublished
		CP000729	17 531
Staphylococcus aureus ssp. aureus JH1	Firmicutes	CP000736	2 906 700	2780	JGI	Unpublished
		CP000737	30 429
Staphylococcus aureus ssp. aureus str. Newman	Firmicutes	AP009351	2 878 897	2614	Juntendo Univ.	Unpublished
Coxiella burnetii Dugway 7E9-12	γ-Proteobacteria	CP000733	2 158 758	2125	JCVI	Unpublished
			54 179
Haemophilus influenzae PittEE	γ-Proteobacteria	CP000671	1 813 033	1623	Allegheny Institute	Unpublished
Haemophilus influenzae PittGG	γ-Proteobacteria	CP000672	1 887 192	1670	Allegheny Institute	Unpublished
Pseudomonas aeruginosa PA7	γ-Proteobacteria	CP000744	6 588 339	6286	JCVI	Unpublished
Shewanella baltica OS185	γ-Proteobacteria	CP000753	5 229 686	4394	JGI	Unpublished
		CP000754	83 224
Yersinia pseudotuberculosis IP 31758	γ-Proteobacteria	CP000720	4 723 306	4324	JCVI	Unpublished
		CP000719	153 140
		CP000718	58 679
Campylobacter jejuni ssp. doylei 269.97	ε-Proteobacteria	CP000768	1 845 106	1731	JCVI	Unpublished

Sequencing centre names are abbreviated as follows: Allegeny Institute, Allegheny-Singer Research Institute, Pittsburgh, PA, USA; CNRS-Marseille, CNRS – UPR2589, Institut de Biologie structurale et Microbiologie, Marseille, France; INRA – Jouy-en-Josas, Unité Virologie et Immunologie Moléculaires, Institut National de la Recherche Agronomique, Jouy-en-Josas, France; JAMSTEC, Japan Agency for Maine-Earth Science and Technology, Natsushima-cho, Yokosuka, Japan; JCVI, J. Craig Venter Institute, Rockville, MD, USA; JGI, US Department of Energy Joint Genome Institute, Walnut Creek, CA, USA; Juntendo Univ., Department of Bacteriology at Juntendo University, Bunkyo-ku, Tokyo, Japan; Los Alamos, Bioscience Division, Los Alamos National Laboratory, Los Alamos, NM, USA; U. Göttingen, Göttingen Genomics Laboratory at the Institute of Microbiology and Genetics, University of Göttingen, Göttingen, Germany; Univ. Tokyo, Department of Biological Sciences, Graduate School of Science, University of Tokyo, Tokyo, Japan; WashU, Washington University School of Medicine, St. Louis, MO, USA.

The TM7 phylum was identified based on the comparison of 16S rRNA genes in samples from a variety of terrestrial (peat bog, activated sludge) and aquatic environments. Fluorescence in situ hybridization revealed cells forming long, thick (up to 50 × 4 μm) filaments (Hugenholtz ). Similar sequences were subsequently detected in deep-sea hydrothermal sediment, humic lake, hypersaline wastewater and even in a marine sponge (Lopez-Garcia ; Lefebvre ; Newton ; Thiel ). Members of TM7 were also detected in human oral cavity and oesophagus, often associated with necrotizing ulcerative gingivitis, halitosis and periodontitis (Paster ; Brinig ; Kazor ; Pei ). Still, no representative of the TM7 was ever obtained in a pure culture. Genome sequencing of TM7 was made possible by capturing individual cells in a specially designed microfluidic device, followed by DNA amplification and sequencing (Marcy ). Although the genome size and the number of encoded proteins could not be reliably estimated, the genome assembly produced a total of 2.86 Mb containing 3245 predicted genes. Some of the predicted genes did not have known homologues, while those that did had relatively low sequence identity to genes from known phyla. These observations further confirmed that the analyzed genes came from a representative of a new phylum. Mapping the predicted genes onto the metabolic pathway map suggested that the TM7 isolate was able to perform glycolysis, the tricarboxylic acid cycle, nucleotide biosynthesis and some amino acid biosyntheses. Obviously, the incomplete genome did not allow identification of the missing pathways that might give some clues to the reasons why members of TM7 refuse to grow in pure culture. Anyway, this work represents a significant step towards characterization of these fascinating organisms.

Another major news was successful transformation of Mycoplasma capricolum cells with full-length chromosomal DNA from Mycoplasma mycoides by a group at the J. C. Venter Institute (Lartigue ). The efficiency of transformation (referred to as ‘genome transplantation’ by the authors) reached one recipient per 150 000 cells. This is a significant technical accomplishment that opens new possibilities for gene manipulation in the framework of the so-called ‘synthetic biology’.

As mentioned above, scientists at the University of Tokyo, Japan, revised the previously reported genomic sequence of the hot-spring red alga C. merolae (Matsuzaki ), filled all of the 46 remaining gaps, sequenced the 34 remaining chromosomal ends, and reported the first eukaryotic nuclear-genome sequence that is 100% complete (Nozaki ). The total genome of C. merolae is 16 728 945 nucleotides (nt) in size and consists of 20 linear chromosomes with a total of 16 546 747 nt, circular chloroplast genome of 149 987 nt, and a circular mitochondrial genome of 32 211 nt. Each chromosome encodes between 102 and 484 proteins, for a total of 4775, the chloroplast DNA encodes 208 and the mitochondrial DNA encodes 34 proteins. The small size of the C. merolae protein set, coupled with the fact that only a small fraction (∼0.5%) of these 5017 genes contain introns, makes this thermophilic (45°C) alga a perfect model organism for studying all kinds of eukaryotic proteins.

For the past 8 years, Deinococcus radiodurans remained the only highly radioresistant bacterium with a completely sequenced genome, which severely limited the use of comparative genomics to analyze the resistance mechanisms. The scientists at the US Department of Energy Joint Genome Institute (JGI) have now completed genome sequencing of another radioresistant bacterium, Kineococcus radiotolerans, which opens the possibility for meaningful genome comparisons. Kineococcus radiotolerans is an aerobic actinobacterium isolated from a highly radioactive waste at the Savannah River Technology Center in Aiken, South Carolina (Phillips ). Like D. radiodurans, this bacterium is highly resistant both to the ionizing γ-radiation and to desiccation. Cells of K. radiotolerans produce an orange carotenoid pigment and move by means of polar flagella. The genome encodes all key components of the chemotaxis machinery, including 22 methyl-accepting chemotaxis sensor proteins, by far the largest number found in any actinobacteria. The high resistance of K. radiotolerans to dessication suggests that related bacteria might be widespread in arid desert environments.

Although human intestine is not typically perceived as a subject of environmental studies, scientists at the Washington University have embarked on an extensive project aimed at characterizing the microbial diversity in the human intestinal tract, the Human Gut Microbiome (see http://genome.wustl.edu/hgm/HGM_frontpage.cgi). In the previous column we have discussed completion of the genome of the intestinal methanogenic archaeon Methanobrevibacter smithii (Samuel ). The same group has now released complete genomes of two members of the phylum Bacteroidetes that are prominently represented in the distal gut of healthy humans, Bacteroides distasonis (recently reclassified as Parabacteroides distasonis) and Bacteroides vulgatus (Xu ). This paper (which is freely available online) describes a detailed comparison of P. distasonis and B. vulgatus genomes with the previously sequenced genomes of two other gut symbionts, Bacteroides fragilis and Bacteroides thetaiotaomicron, and analyzes the role of lateral gene transfer and gene duplication in the adaptation of Bacteroides spp. to the gut environment.

One more representative of the Bacteroidetes is the widespread fish pathogen Flavobacterium psychrophilum, whose genome description (Duchaud ) was published a month after the release of the genome sequence of the closely related soil bacterium Flavobacterium johnsoniae. Flavobacterium psychrophilum infects salmon and trout causing haemorrhagic septicaemia, referred to as ‘rainbow trout fry syndrome’, in young fish and severe necrotic lesions called ‘cold-water disease’ in adult fish. In accordance with its name, F. psychrophilum grows best at 15°C and is most deadly at temperatures in the 3–15°C range. This bacterium is capable of forming biofilms that can survive in stream water for several months. Accordingly, the genome sequence revealed a cluster of genes involved in the biosynthesis of exopolysaccharides. It also encodes a protein similar to cyanophicin synthase, suggesting that F. psychrophilum is capable of storing amino acids, which could contribute to its long-term survival outside of the fish host. The F. psychrophilum genome encodes a number of virulence factors, including various proteases, cytolytic toxins and adhesive proteins. Analysis of these virulence factors and other surface proteins should help identify potential vaccine candidates to protect farmed salmon and rainbow trout against infection by F. psychrophilum.

The current list (Table 1) includes 10 genomes of low G+C Gram-positive bacteria (Firmicutes), two of which represent the genus Bacillus, six come from the family Clostridiaceae (including three environmental isolates and three new strains of the food-borne pathogen Clostridium botulinum) and the remaining two come from new strains of the opportunistic pathogen Staphylococcus aureus.

Bacillus amyloliquefaciens is an soil bacterium that often colonizes plant rhizosphere, promoting plant growth and suppressing plant pathogens. The plant growth-promoting effect has been attributed to the extracellular phytase activity (degradation of inositol hexaphosphate), which provides the plant with phosphate (Idriss ). In addition to phytase, B. amyloliquefaciens secretes numerous amylases, glucanases and proteases, as well as antibacterial and antifungal compounds. The sequenced strain FZB42 encodes several polyketide synthases, two of which has been shown to be responsible for the synthesis of the polyketide antiobiotics bacillaene and difficidin (Chen ).

Bacillus cereus ssp. cytotoxis is a food-borne pathogen, whose genome revealed a significant degree of divergence from the typical B. cereus (Lapidus ). Based on these comparisons, the authors suggest that it should be reclassified as a new species Bacillus cytotoxicus.

Alkaliphilus metalliredigens, a member of the family Clostridiaceae, has been isolated from leachate ponds at the US Borax company in Boron, California, using an enrichment for the ability to reduce Fe(III) in anaerobic conditions at alkaline pH values (Ye ). Alkaliphilus metalliredigens is a strict anaerobe that could tolerate up to 1.5% sodium tetraborate (Na2B4O7) and grew using Fe(III)-citrate, Fe(III)-EDTA, Co(III)-EDTA or Cr(VI) as electron acceptors; yeast extract or lactate served as electron donors. Growth during iron reduction occurred over the pH range of 7.5–11.0 with optimum at pH 9.5, at temperatures ranging from 4°C to 45°C. These properties make A. metalliredigens an attractive candidate for bioremediation of metal-contaminated alkaline environments.

Interestingly, another anaerobic iron-reducing bacterium with a recently sequenced genomes belongs to an entirely different phylogenetic lineage, the δ-Proteobacteria. Anaeromyxobacter strain Fw109-5 has been isolated from an uranium-contaminated subsurface sediment in Oak Ridge, Tennessee (van Landschoot and de Ley, 1983). Although it is an anaerobe, it tolerates microaerophilic conditions and uses acetate, lactate and pyruvate as electron donors and Fe(III) or nitrate as electron acceptors.

Clostridium beijerinckii strain NCIMB 8052 is also a strict anaerobe of potential use in biotechnology. It is a soil isolate that ferments a wide range of carbohydrates (pentoses, hexoses, starch and others) to acetate, butyrate, lactate and other products, including valuable solvents acetone, butanol and isopropanol. Analysis of C. beijerinckii genome and its comparison with the genome of the closely related solventogenic bacterium Clostridium acetobutylicum is expected to provide insight into the mechanisms of solventogenesis and pave way to designing more efficient producers of acetone and butanol suitable for industrial use.

The γ-proteobacterium Actinobacillus succinogenes, isolated from the bovine rumen (Guettler ), is yet another microorganism with potential use in biotechnology. It metabolizes a wide range of sugars (including glucose, fructose, xylose, lactose, and cellobiose), producing succinate, which is a precursor for a number of useful chemical compounds (Zeikus ).

The γ-proteobacterium Klebsiella pneumoniae is best known as an opportunistic human pathogen that causes pneumonia and urinary tract infections in hospital settings and in immunocompromised patients. However, it is a widespread environmental organism, commonly found in soil and water habitats. Distinctive features of Klebsiella cells include an extracellular polysaccharide capsule and the ability to fix nitrogen. The sequenced genome comes from a multiple antibiotic-resistant strain Klebsiella pneumoniae ssp. pneumoniae MGH 78578 that was isolated in 1994 from a pneumonia patient.

The γ-proteobacterium Marinomonas sp. strain MWYL1 was isolated near the North Norfolk, England, from the root surface of the salt marsh grass Spartina anglica. This grass, as well as some microalgae, produces the osmoprotective compound dimethylsulfoniopropionate, which Marinomonas sp. MWYL1 can use as sole carbon source. Metabolism of dimethylsulfoniopropionate produces dimethylsulfide, which is released into the air (Ansede ) and represents a major contribution to sulfur cycling in the marine environment. Products of dimethylsulfide oxidation in the atmosphere act as cloud condensation nuclei and are largely responsible for forming the cloud cover over the oceans, affecting the climate wordwide (Simó, 2001). The mechanism of dimethylsulfide formation was recently resolved (Todd ); genome analysis of Marinomonas MWYL1 could clarify the regulation of this process.

The four sequenced members of the ε-subdivision of the Proteobacteria nicely represent the diversity of this group. Campylobacter curvus and Campylobacter hominis are gastric pathogens closely related to the better-known Campylobacter jejuni, whose genome was recently re-annotated (Gundogdu ). In contrast, Nitratiruptor sp. strain SB155-2 and Sulfurovum sp. strain NBC37-1 have been isolated from the deep-sea vents in the Iheya North hydrothermal field, Japan. These bacteria are chemolithoautotrophs that use hydrogen, sulfide, elemental sulfur or thiosulfate as electron donors and oxygen or nitrate as electron acceptors. They are representative of the microbial ‘dark matter’ in the vicinity of the vents, where ε-proteobacteria comprise a significant fraction of the total microbial population (Nakagawa ). Surprisingly, genome comparisons showed that vent bacteria share with pathogenic ε-proteobacteria a number of genes that had been previously identified as virulence factors (Nakagawa ). These include genes responsible for N-linked glycosylation, hydrogenase and several other genes. The authors suggest that Campylobacter- and Helicobacter-like pathogens evolved from free-living ε-proteobacteria, similar to Nitratiruptor sp. and Sulfurovum sp.

For the past several years, the early branching bacterial phylum Thermotogales was represented by a single complete genome of Thermotoga maritima (Nelson ). With an increased focus on microbial diversity, JGI has recently launched a new project aimed at obtaining genome sequences of seven more representatives of this interesting phylum. The genome of Thermotoga petrophila, the first one generated by this project, was released earlier this year. The JGI has now released genomes of two more members of the Thermotogales, Fervidobacterium nodosum strain Rt17-B1, isolated from a hot spring in New Zealand, and Thermosipho melanesiensis strain BI429, which was isolated from the gills of a deep-sea vent hydrothermal mussel, Bathymodiolus brevior, from the Lau Basin in the South-western Pacific Ocean (Antoine ). Comparison of the genomes of hot-spring and marine isolates of Thermotogales is expected to shed light on the mechanisms of survival in high-pressure marine environments and allow re-assessing the degree of lateral gene transfer from archaea, which in T. maritima was estimated to reach 20% of all genes (Nelson ).

In other genomics news, Jon Hobman, Charles Penn and Mark Pallen of the University of Birmingham have stirred the pot by publishing a paper with the provocative title ‘Laboratory strains of Escherichia coli: model citizens or deceitful delinquents growing old disgracefully?’, which states, inter alia, that ‘that microbiology's chief idol has feet of clay’ (Hobman ). Despite somewhat hyped rhetoric, this paper makes a number of valid points, mentioning that E. coli K-12 has undergone numerous passages on rich media and cycles of mutagenesis and is hardly representative of either the ancestral E. coli or the current environmental and ‘enteropathogenic, enterotoxigenic, enteroinvasive, enterohaemorrhagic, enteroaggregative and diffusely adherent’ strains. In what might be particularly relevant to the subject of this journal, the authors warn against ‘assuming that any models of global regulation or metabolic flux can be generalized to E. coli in a state of nature’, look forward to obtaining hundreds, if not thousands, of genome sequences of naturally occurring relatives of E. coli K-12 and welcome ‘the bright new, pluralist, genome-saturated “eco-evo” future of E. coli’. Thus, aside from the title (and section subtitles), there seems to be very little in this paper to argue about. However, from the genome analysis point of view, it appears that the authors have overlooked a major drawback in our description(s) of E. coli K-12, namely the fact that at least one-third of its genes still have unknown (or poorly characterized) function (Riley ). For example, considering the very similar sets of signal transduction proteins encoded in E. coli K-12 and in all other E. coli genomes sequenced to date, one has to conclude that we still have only a vague understanding of the functions of its 30 histidine kinases, 29 diguanylate cyclases and/or c-di-GMP-specific phosphodiesterases and two predicted Ser/Thr protein kinases (M.Y. Galperin, in preparation). Escherichia coli K-12 still represents our best hope to achieve a complete understanding of the genome of a free-living bacterium and deserves to be treated as such.

In addition to the deliberately provocative comment on E. coli, Mark Pallen got involved in another controversy, this time through no fault of his own. About a year ago, Pallen teamed up with Nick Matzke, an evolutionary biologist at the National Center for Science Education in Oakland, California (http://www.natcenscied.org/), to produce a wide-ranging analysis of the evolution of bacterial flagella and refute the claims of proponents of the ‘intelligent design’ on the ‘irreducible complexity’ of that organelle [Pallen and Matzke, 2006; see also the Panda's Thumb weblog (http://www.pandasthumb.org/) and the paper by Scott and Matzke (2007) on the history of the ‘intelligent design’ movement]. In April 2007, Renyi Liu and Howard Ochman also published a paper (freely available online) aimed at refuting the ‘intelligent design’ views on flagellar origin. Liu and Ochman (2007a) compared flagellar proteins from various bacterial genomes using pairwise blast searches with the blast2seq program (Tatusova and Madden, 1999) and detected a certain degree of sequence similarity between nearly all types of proteins. Although in many cases the similarity levels were not statistically significant (unless the low-complexity filtering was deliberately switched off), Liu and Ochman interpreted their results as an evidence of common origin (= homology) between all flagellar proteins, even those that had been known to have different three-dimensional structures and were obviously non-homologous. Those shaky blast results were presented in a form of a ‘network of relationships among flagellar core proteins’, which conveyed an aura of infallibility that must have swayed gullible reviewers and editors of PNAS. This paper attracted a positive comment in ScienceNOW (Cutraro, 2007) but was met with a barrage of criticism, including numerous postings on the Panda's Thumb website and T. taxus blog (http://ttaxus.blogspot.com/2007/05/jcvi-evolutionary-genomics-journal-club.html) suggesting that what is true in the paper by Liu and Ochman (2007a) is not new, and what is new is not true. In addition, a recent paper by Doolittle and Zhaxybayeva (2007) questioned the validity of the phylogenetic analysis in that paper. In response, Liu and Ochman published a correction, admitting switching off the low-complexity filter and using a more permissive 9 × 10−4 cut-off E-value instead of the 10−4 value given in the original publication, but claiming that ‘These errors do not affect the conclusions of the article’ (Liu and Ochman, 2007b). This story is interesting not only because PNAS has published a deeply flawed paper, something that has happened previously in other prestigious journals, particularly when dealing with ‘hot’ topics. Rather, this case illustrates the caveats of automated sequence analysis, which can only be trusted if the results pass a ‘sanity check’ by a well-trained biologist. Unfortunately, perfunctory sequence analysis has already caused a number of major blunders (Iyer ) and is likely to generate many more. [Full disclosure: the author was the editor of the original blast2seq paper (Tatusova and Madden, 1999) and is one of the authors of a paper on the possible origin of flagellar ATPases (Mulkidjanian )].

Finally, Minoru Kanehisa and colleagues at the University of Kyoto reported an analysis of 191 completely sequenced genomes, aimed at answering a key question: are there any additional directly encoded unusual amino acids besides selenocysteine and pyrrolysine? As these two amino acids (respectively, the 21st and 22nd ones) are both encoded by stop codons, the genomes were inspected for the conservation patterns in the vicinity of the predicted stop codons (Fujita ). This search failed to find new conserved contexts, which suggested that the 23rd amino acid either has a very limited phylogenetic distribution or does not exist at all.