Open Access

Genome analysis of Desulfotomaculum kuznetsovii strain 17T reveals a physiological similarity with Pelotomaculum thermopropionicum strain SIT

  • Michael Visser1,
  • Petra Worm1,
  • Gerard Muyzer2,
  • Inês A. C. Pereira3,
  • Peter J. Schaap4,
  • Caroline M. Plugge1,
  • Jan Kuever5,
  • Sofiya N. Parshina6,
  • Tamara N. Nazina6,
  • Anna E. Ivanova6,
  • Rizlan Bernier-Latmani7,
  • Lynne A. Goodwin8, 9,
  • Nikos C. Kyrpides8,
  • Tanja Woyke8,
  • Patrick Chain8, 9,
  • Karen W. Davenport8, 9,
  • Stefan Spring10,
  • Hans-Peter Klenk10 and
  • Alfons J. M. Stams1, 11
Standards in Genomic Sciences20138:8010069

DOI: 10.4056/sigs.3627141

Published: 15 April 2013

Abstract

Desulfotomaculum kuznetsovii is a moderately thermophilic member of the polyphyletic spore-forming genus Desulfotomaculum in the family Peptococcaceae. This species is of interest because it originates from deep subsurface thermal mineral water at a depth of about 3,000 m. D. kuznetsovii is a rather versatile bacterium as it can grow with a large variety of organic substrates, including short-chain and long-chain fatty acids, which are degraded completely to carbon dioxide coupled to the reduction of sulfate. It can grow methylotrophically with methanol and sulfate and autotrophically with H2 + CO2 and sulfate. For growth it does not require any vitamins. Here, we describe the features of D. kuznetsovii together with the genome sequence and annotation. The chromosome has 3,601,386 bp organized in one contig. A total of 3,567 candidate protein-encoding genes and 58 RNA genes were identified. Genes of the acetyl-CoA pathway, possibly involved in heterotrophic growth with acetate and methanol, and in CO2 fixation during autotrophic growth are present. Genomic comparison revealed that D. kuznetsovii shows a high similarity with Pelotomaculum thermopropionicum. Genes involved in propionate metabolism of these two strains show a strong similarity. However, main differences are found in genes involved in the electron acceptor metabolism.

Keywords

Thermophilic spore-forming anaerobes sulfate reduction autotrophic methylotrophic Peptococcaceae Clostridiales

Introduction

Desulfotomaculum kuznetsovii strain 17T (VKM B-1805; DSM 6115) is a moderately thermophilic sulfate-reducing bacterium isolated from deep subsurface thermal mineral water [1]. It grows with a wide range of substrates, including organic acids, such as long-chain fatty acids, short-chain fatty acids (butyrate, propionate, acetate), lactate, pyruvate, fumarate and succinate as well as ethanol and methanol. These substrates are degraded to CO2 coupled to sulfate reduction. The strain is also able to grow autotrophically with H2/CO2 and sulfate and to ferment pyruvate and fumarate. For growth, D. kuznetsovii has no vitamin requirement.

Desulfotomaculum is a genus of Gram-positive, spore-forming anaerobes that is phylogenetically and physiologically very diverse. The genus is poorly studied physiologically, while its members are known to play an important role in the carbon and sulfur cycle in a variety of often adverse environments. The genus is divided phylogenetically into different sub-groups [2,3]. To get a thorough understanding of the evolutionary relationship of the different Desulfotomaculum sub-groups and the physiology of the individual species, it is important to have genome sequence information. Here, we present a summary of the features of D. kuznetsovii strain 17T, together with the description of the complete genomic sequencing and annotation. Moreover, we describe a physiological and genomic comparison of D. kuznetsovii strain 17T and Pelotomaculum thermopropionicum strain SIT, because phylogenetically P. thermopropionicum is the closest related organism with validly published name that has a completely sequenced genome. However, the two strains have different physiological traits. For example, P. thermopropionicum is not able to grow by sulfate reduction, but is able to grow in syntrophy with methanogens. D. kuznetsovii lacks this ability. By comparing the genomes of the two bacteria we were able to identify the main similarities and differences.

Classification and features

D. kuznetsovii is a member of the phylum Firmicutes. Phylogenetic analysis of the 16S rRNA genes of D. kuznetsovii shows that it clusters in Desulfotomaculum cluster 1. This cluster not only contains Desulfotomaculum species, but also members of the genera Sporotomaculum, Cryptanaerobacter and Pelotomaculum. D. kuznetsovii is part of sub-group 1c together with D. solfataricum, D. luciae, D. thermosubterraneum, D. salinum, D. australicum, and D. thermocisternum, while Pelotomaculum species belong to sub-group 1h (Figure 1) [2].
Figure 1.

Neighbor joining tree based on 16S rRNA sequences showing the phylogenetic affiliation of Desulfotomaculum and related species divided in the subgroups of Desulfotomaculum cluster 1. D. kuznetsovii is printed in bold type. The sequences of different Thermotogales were used as outgroup, but were pruned from the tree. Closed circles represent bootstrap values between 75 and 100%. The scale bar represents 10% sequences difference.

D. kuznetsovii cells are rod-shaped (1.0–1.4 x 3.5–5 εm) with rounded ends and peritrichous flagella [Figure 2]. Spores of D. kuznetsovii are spherical (1.3 εm in diameter) and centrally located causing swelling of the cells. D. kuznetsovii grows between 50 and 85°C, but the optimal growth temperature is 60–65°C. The substrates D. kuznetsovii can grow with are completely oxidized to CO2. Suitable electron acceptors are sulfate, thiosulfate and sulfite. D. kuznetsovii is also able to grow by fermentation of pyruvate and fumarate. A summary of the classification and general features of D. kuznetsovii is presented in Table 1 [1].
Figure 2.

Scanning electron microscopic photograph of D. kuznetsovii.

Table 1.

Classification and general features of D. kuznetsovii DSM 6115 according to the MIGS recommendations [4].

MIGS ID

Property

Term

Evidence codea

 

Current classification

Domain Bacteria

TAS [5]

 

Phylum Firmicutes

TAS [68]

 

Class Clostridia

TAS [9,10]

 

Order Clostridiales

TAS [11,12]

 

Family Peptococcaceae

TAS [11,13]

 

Genus Desulfotomaculum

TAS [11,14,15]

 

Species Desulfotomaculum kuznetsovii

TAS [1,16]

 

Type strain 17

 
 

Gram stain

Positive

TAS [1]

 

Cell shape

Rods

TAS [1]

 

Motility

peritrichous flagella

TAS [1]

 

Sporulation

oval, terminal or subterminal, slightly swelling the cell.

TAS [1]

 

Temperature range

50–85°C

TAS [1]

 

Optimum temperature

60–65°C

TAS [1]

 

Carbon source

CO2 (autotrophic) and organic substrates (heterotrophic)

TAS [1]

 

Energy source

Sulfate-dependent growth and fermentative growth with pyruvate and fumarate.

TAS [1]

 

Electron acceptor

Sulfate, thiosulfate and sulfite.

TAS [1]

MIGS-6

Habitat

Geothermal groundwater, sediment and hot solfataric fields.

TAS [1,17,18]

MIGS-6.3

Salinity

2–3% NaCl

TAS [1]

MIGS-22

Oxygen

Obligate anaerobes

TAS [1]

MIGS-15

Biotic relationship

Free living

TAS [1]

MIGS-14

Pathogenicity

None

 

MIGS-4

Geographic location

Sukhumi, Georgia

TAS [1]

MIGS-5

Sample collection time

1987 or before

TAS [1]

MIGS-4.1

Latitude

43.009

TAS [1]

MIGS-4.2

Longitude

40.989

TAS [1]

MIGS-4.3

Depth

2800–3250 m

TAS [1]

Evidence codes - TAS: Traceable Author Statement (i.e., a direct report exists in the literature); NAS: Non-traceable Author Statement (i.e., not directly observed for the living, isolated sample, but based on a generally accepted property for the species, or anecdotal evidence). Evidence codes are from the Gene Ontology project [19].

Genome sequencing and annotation

Genome project history

D. kuznetsovii was selected for sequencing in the DOE Joint Genome Institute Community Sequencing Program 2009, proposal 300132_795700 ‘Exploring the genetic and physiological diversity of Desulfotomaculum species’, because of its phylogenetic position in one of the Desulfotomaculum sub-groups, its important role in bioremediation, and its ability to use propionate, acetate and methanol for growth. The genome project is listed in the Genome OnLine Database (GOLD) [20] as project Gc01781, and the complete genome sequence was deposited in Genbank. Sequencing, finishing and annotation of the D. kuznetsovii genome were performed by the DOE Joint Genome Institute (JGI). A summary of the project information is shown in Table 2.
Table 2.

Genome sequencing project information

MIGS ID

Property

Term

MIGS-31

Finishing quality

Finished

MIGS-28

Libraries used

Four genomic libraries: one Illumina shotgun library, one 454 standard library, two paired end 454 libraries

MIGS-29

Sequencing platforms

Illumina GAii, 454 Titanium

MIGS-31.2

Fold coverage

158.2 × illumina; 30.6 × pyrosequencing

MIGS-30

Assemblers

VELVET version 0.7.63; Newbler version 2.3; phrap version SPS - 4.24

MIGS-32

Gene calling method

Prodigal 1.4, GenePRIMP

 

INSDC ID

CP002770.1

 

Genome Database release

July 20, 2012

 

Genbank Date of Release

May 24, 2011

 

GOLD ID

Gc01781

 

NCBI project ID

48313

MIGS-13

Source material identifier

DSM 6115T

 

Project relevance

Obtain insight into the phylogenetic and physiological diversity of Desulfotomacum species, and bioremediation.

Growth conditions and DNA isolation

D. kuznetsovii was grown anaerobically at 60oC in bicarbonate buffered medium with propionate and sulfate as substrates [1]. DNA of cell pellets was isolated using the standard DOE-JGI CTAB method recommended by the DOE Joint Genome Institute (JGI, Walnut Creek, CA, USA). In short, cells were resuspended in TE (10 mM tris; 1 mM EDTA, pH 8.0). Subsequently, cells were lysed using lysozyme and proteinase K, and DNA was extracted and purified using CTAB and phenol:chloroform:isoamylalcohol extractions. After precipitation in 2-propanol and washing in 70% ethanol, the DNA was resuspended in TE containing RNase. Following a quality and quantity check using agarose gel electrophoresis in the presence of ethidium bromide, and spectrophotometric measurement using a NanoDrop ND-1000 spectrophotometer (NanoDrop® Technologies, Wilmington, DE, USA).

Genome sequencing and assembly

The genome was sequenced using a combination of Illumina and 454 sequencing platforms. All general aspects of library construction and sequencing can be found at the JGI website [21]. Pyrosequencing reads were assembled using the Newbler assembler (Roche).

The initial Newbler assembly consisting of 81 contigs in five scaffolds was converted into a phrap [22] assembly by making fake reads from the consensus, to collect the read pairs in the 454 paired end library. Illumina GAii sequencing data (570.2 Mb) was assembled with Velvet [23] and the consensus sequences were shredded into 1.5 kb overlapped fake reads and assembled together with the 454 data. The 454 draft assembly was based on 134.6 Mb 454 draft data and all of the 454 paired end data. Newbler parameters are -consed -a 50 -l 350 -g -m -ml 20. The Phred/Phrap/Consed software package [22] was used for sequence assembly and quality assessment in the subsequent finishing process. After the shotgun stage, reads were assembled with parallel phrap (High Performance Software, LLC). Possible mis-assemblies were corrected with gapResolution [21], Dupfinisher [24], or sequencing cloned bridging PCR fragments with subcloning. Gaps between contigs were closed by editing in Consed, by PCR and by Bubble PCR primer walks (J.-F. Chang, unpublished). A total of 400 additional reactions and one shatter library were necessary to close gaps and to raise the quality of the finished sequence. Illumina reads were also used to correct potential base errors and increase consensus quality using a software Polisher developed at JGI [25]. The error rate of the completed genome sequence is less than 1 in 100,000. Together, the combination of the Illumina and 454 sequencing platforms provided 188.8 × coverage of the genome. The final assembly contained 323,815 pyrosequence and 15,594,144 Illumina reads.

Genome annotation

Genes were identified using Prodigal [26] as part of the Oak Ridge National Laboratory genome annotation pipeline, followed by a round of manual curation using the JGI GenePRIMP pipeline [27]. The predicted CDSs were translated and used to search the National Center for Biotechnology Information (NCBI) nonredundant database, UniProt, TIGR-Fam, Pfam, PRIAM, KEGG, COG, and InterPro databases. Additional gene prediction analysis and functional annotation was performed within the Integrated Microbial Genomes - Expert Review (IMG-ER) platform [28].

Genome properties and genome comparison with other strains

The genome of D. kuznetsovii consists of a circular chromosome of 3,601,386 bp with 54.88% GC content (Table 3 and Figure 3). Pseudogenes comprise 4.66% of the genes identified. Of the 3,625 genes predicted, 3,567 are protein-coding genes of which 2,560 are assigned to COG functional categories. The distribution of these genes into COG functional categories is presented in Table 4.
Figure 3.

Graphical map of the chromosome of D. kuznetsovii. From outside to the center: Genes on the forward strand (colored by COG categories), Genes on the reverse strand (colored by COG categories), RNA genes (tRNAs green, rRNAs red, other RNAs black), GC content, GC skew.

Table 3.

Genome statistics

Attribute

Value

% of totala

Genome size (bp)

3,601,386

100.00

Genome coding region (bp)

3,057,959

84.91

Genome G+C content (bp)

1,976,601

54.88

Total genes

3,625

100.00

RNA genes

58

1.60

Protein-coding genes

3,567

98.40

Genes in paralog clusters

1,373

37.88

Genes assigned to COGs

2,560

70.62

Pseudo genes

169

4.66

Genes with signal peptides

582

16.06

Genes with transmembrane helices

748

20.63

Table 4.

Number of genes associated with the general COG functional categories

Code

Value

%agea

Description

J

148

5.32

Translation

A

0

0.00

RNA processing and modification

K

184

6.61

Transcription

L

207

7.44

Replication, recombination and repair

B

2

0.07

Chromatin structure and dynamics

D

60

2.16

Cell cycle control, mitosis and meiosis

Y

0

0.00

Nuclear structure

V

35

1.26

Defense mechanisms

T

177

6.36

Signal transduction mechanisms

M

122

4.38

Cell wall/membrane biogenesis

N

79

2.84

Cell motility

Z

2

0.07

Cytoskeleton

W

0

0.00

Extracellular structures

U

75

2.69

Intracellular trafficking and secretion

O

81

2.91

Posttranslational modification, protein turnover, chaperones

C

261

9.38

Energy production and conversion

G

106

3.81

Carbohydrate transport and metabolism

E

197

7.08

Amino acid transport and metabolism

F

55

1.98

Nucleotide transport and metabolism

H

158

5.68

Coenzyme transport and metabolism

I

89

3.20

Lipid transport and metabolism

P

127

4.56

Inorganic ion transport and metabolism

Q

122

3.58

Secondary metabolites biosynthesis, transport and catabolism

R

331

11.89

General function prediction only

S

257

9.23

Function unknown

-

1,065

29.38

Not in COGs

The genome of D. kuznetsovii has 58 RNA genes of which, three are 16S rRNA genes. This is one more than the previously described rrnA and rrnB [29]. These two rRNA genes contained two large inserts. One at the variable 5′terminal region and one at the variable 3′terminal region. The main differences between the two rRNA genes were found in these inserts. These inserts were hypothesized to be involved in the operation of ribosomes at high temperatures. However, more research is needed to assess the function of these inserts. All three rRNA genes of D. kuznetsovii have a size of approximately 1,700 nucleotides. This suggests that the third rRNA gene might also contain inserts. Alignment of the 16S rRNA genes confirmed the presence of inserts in all three 16S rRNA genes (data not shown).

BLAST analysis [30,31] of the genes of D. kuznetsovii against genes in the KEGG Sequence Similarity DataBase revealed similarity with other Desulfotomaculum strains (Table 5), D. acetoxidans, D. carboxydivorans, “D. reducens” and D. ruminis, but interestingly also with non-Desulfotomaculum strains. D. kuznetsovii contains 873 genes with high similarity to genes of Pelotomaculum thermopropionicum, which is more than to any of the sequenced Desulfotomaculum species. Moreover, we identified the conserved proteins of D. kuznetsovii across three related fully sequenced species (Table 6). The bidirectional best blast hits showed that despite the smaller genome of P. thermopropionicum it contained more homologous predicted proteins with D. kuznetsovii (1,406) compared to D. acetoxidans (1,309) and “D. reducens” (1330). This suggests a strong physiological similarity between D. kuznetsovii and P. thermopropionicum.
Table 5.

Taxonomic distribution of the top KEGG hits of D. kuznetsovii genes based on BLAST against KEGG database.

Kingdom

Category

Species

Hits

Archaea

  

91

 

Crenarchaeota

 

9

 

Euryarchaeota

 

81

 

Thaumarchaeota

 

1

Bacteria

  

2,963

 

Acidobacteria

 

2

 

Actinobacteria

 

16

 

Alphaproteobacteria

 

13

 

Bacteroidetes

 

5

 

Betaproteobacteria

 

14

 

Cyanobacteria

 

19

 

Deinococcus-Thermus

 

16

 

Deltaproteobacteria

 

62

 

Epsilonproteobacteria

 

1

 

Firmicutes

 

2,728

  

Ammonifex degensii

170

  

Carboxydothermus hydrogenoformans

58

  

Desulfotomaculum acetoxidans

310

  

Candidatus Desulforudis audaxviator

154

  

Desulfotomaculum carboxydivorans

268

  

Desulfotomaculum reducens

111

  

Desulfotomaculum ruminis

132

  

Moorella thermoacetica

183

  

Pelotomaculum thermopropionicum

873

  

Thermincola potens JR

104

 

Fusobacteria

 

2

 

Gammaproteobacteria

 

12

 

Green nonsulfur bacteria

 

20

 

Green sulfur bacteria

 

4

 

Hyperthermophilic bacteria

 

31

 

OtherProteobacteria

 

1

 

Spirochaetes

 

9

 

Synergistetes

 

6

 

Verrucomicrobia

 

2

Eukaryotes

  

3

 

Plants

 

1

 

Protists

 

2

Null

  

342

Total

  

3,399

Species that had more than 50 genes similar to D. kuznetsovii were included in this table, others were only summarized in categories.

Table 6.

Proteins of D. kuznetsovii conserved across three related species with fully sequenced genomes.

Subject DB Input Query

D. acetoxidans

D. kuznetsovii

D. reducens

P. thermopropionicum

D. acetoxidans

4,068

1,539

1,525

1,486

1,309

1,316

1,255

D. kuznetsovii

1,509

 

1,518

1,645

 

1,309

3,398

1,330

1,406

D. reducens

1,537

1,571

 

1,438

 

1,316

1,330

3,276

1,211

P. thermopropionicum

1,430

1,600

1,395

 
 

1,255

1,406

1,211

2,919

BLAST analyses were performed using standard settings and best hits were filtered for 40% identity over an alignment length of 75 amino acids as a minimum requirement. The values show the number of predicted proteins that are homologous to the query species in each row. The number of similar proteins obtained with a unidirectional BLAST is indicated in light blue. Bidirectional best blast hits are indicated in dark blue. Proteomes were obtained from ftp.ncbi.nih.gov/Bacteria/. Accession numbers are in parenthesis: Desulfotomaculum acetoxidans (NC_013216); Desulfotomaculum kuznetsovii (NC_015573); “Desulfotomaculum reducens” (NC_009253); Pelotomaculum thermopropionicum (NC_009454).

Insights into the genome

Involvement of the acetyl-coA pathway in growth with acetate and methanol

D. kuznetsovii oxidizes acetate completely to CO2. The pathway of acetate degradation has not been studied yet, but sulfate reducers may employ the tricarboxylic acid (TCA) cycle or the acetyl-CoA pathway for acetate degradation, as exemplified by Desulfobacter postgatei and Desulfobacca acetoxidans, respectively [32]. Most genes predicted to code for enzymes of the TCA cycle are present in the genome of D. kuznetsovii, but genes with similarity to those coding for an ATP-dependent citrate synthase and isocitrate dehydrogenase are missing. This suggests that the TCA cycle is not complete and that the TCA cycle enzymes have mainly an anabolic function or a function in other catabolic pathways, such as the propionate degradation pathway. Genes with similarity to those coding for enzymes involved in the acetyl-CoA pathway are all present in the genome of D. kuznetsovii (Figure 4), which suggests its involvement in acetate oxidation. However, there are no genes similar to those that code for acetate kinase and phosphate acetyltransferase present in the genome. The reaction from acetate to acetyl-CoA is likely performed by acetyl-CoA synthetase (Desku_1241).
Figure 4.

Pathway of acetate oxidation to CO2 by D. kuznetsovii. Enzymes in this figure are in bold italic and their locus tags are included. Genes with the locus tags Desku_1488 and Desku_1490 putatively code for the small subunit and the large subunit of the iron-sulfur protein, respectively. This protein is involved in transferring the methyl from acetyl-CoA to tetrahydrofolate. Abbreviations: A-CoA S, acetyl-CoA synthetase; AcsA, carbon-monoxide dehydrogenase; AcsB, acetyl-CoA synthase; CFeSP, iron-sulfur protein; CH3, methyl; THF, tetrahydrofolate; MeTr, methyltransferase.

D. acetoxidans is an acetate-oxidizing Desulfotomaculum species, positioned in sub-group 1e (Figure 1), that also uses the acetyl-CoA pathway for acetate oxidation to CO2 [33]. The genes involved in acetate oxidation in D. acetoxidans are similar to those in D. kuznetsovii, but there are some exceptions. The genome of D. acetoxidans does not contain a gene that putatively codes for acetyl-CoA synthetase, similar to D. kuznetsovii, but contains genes that putatively code for an acetate kinase and a phosphate acetyltransferase [34]. Additionally, putative carbon-monoxide dehydrogenase complex coding genes involved in the acetyl-CoA pathway show differences between the two Desulfotomaculum species. D. kuznetsovii lacks a ferredoxin coding gene that is located between cooC (Desku_1493) and acsE (Desku_1487), which in contrast is present in the genome of D. acetoxidans (Dtox_1273). Moreover, three genes similar to heterodisulfide reductase encoding genes (Desku_1486-1484) are located upstream of acsE in D. kuznetsovii, which is not the case in the genome of D. acetoxidans.

Methanol metabolism

Growth of D. kuznetsovii with methanol and sulfate was studied [35]. In that study the activity of methyltransferase, an enzyme that is involved in methanol metabolism in methanogens and acetogens [36,37], could not be assessed, while low activities of an alcohol dehydrogenase could be measured. An alcohol dehydrogenase with a molecular mass of 42 kDa was partially purified and showed activity with methanol [35]. The genome of D. kuznetsovii contains several alcohol dehydrogenase genes (Desku_0165, 0619, 0624, 0628, 2955, 3082) that each code for an enzyme with a size of approximately 42 kDa. In the genome, genes with similarity to those coding for a methanol methyltransferase mtaA (Desku_0050, 0055, 0060), mtaB (Desku_0051) and mtaC (Desku_0048, 0049, 0052, 0056) were also found, suggesting a methanol metabolism as described in Moorella thermoacetica [36]. Further studies are needed to obtain information about the diversity of the methanol-degradation pathways in D. kuznetsovii.

Comparison of D. kuznetsovii and P. thermopropionicum genomes

Genomic comparison revealed that a large number of D. kuznetsovii genes show similarity to genes of Pelotomaculum thermopropionicum, a syntrophic propionate-oxidizing thermophile (Table 5 and 6). Interestingly, among them are genes that putatively code for enzymes involved in propionate metabolism (Table 7). Moreover, the genetic organization of the methylmalonyl-CoA (mmc) cluster in the genome of both bacteria is similar (Figure 5). However, D. kuznetsovii lacks tps, mmcA and mmcM in the mmc cluster. mmcA codes for a response regulator and mmcM for pyruvate ferredoxin oxidoreductase.
Figure 5.

Gene organization of the mmc cluster in D. kuznetsovii and P. thermopropionicum. Names of the genes can be found in table 6, except for tps, which is a transposase gene.

Table 7.

Genes in D. kuznetsovii that are annotated as enzymes involved in propionate metabolism.

Gene symbol

Locus tag

Function

Homologous protein in P. thermopropionicum

   

Identity (%)

Locus tag

sdhB

Desku_0434

Succinate dehydrogenase, FeS protein

76

PTH_1018

sdhA

Desku_0435

Succinate dehydrogenase, flavoprotein

76

PTH_1017

sdhC

Desku_0436

Succinate dehydrogenase, cytochrome b

51

PTH_1016

citE

Desku_1348

Citrate lyase

57

PTH_1335

sdhA

Desku_1353

Succinate dehydrogenase, flavoprotein

83

PTH_1491

sdhB

Desku_1354

Succinate dehydrogenase, FeS protein

75

PTH_1490

mmcB

Desku_1358

Fumarase, N-terminal domain

73

PTH_1356

mmcC

Desku_1359

Fumarase, C-terminal domain

77

PTH_1357

mmcD2

Desku_1361

Succinyl-CoA synthetase, alpha subunit

78

PTH_1359

mmcE

Desku_1362

Methylmalonyl-CoA mutase, N-terminal domain

77

PTH_1361

mmcF

Desku_1363

Methylmalonyl-CoA mutase, C-terminal domain

82

PTH_1362

mmcG

Desku_1364

Methylmalonyl-CoA epimerase

86

PTH_1363

mmcH

Desku_1365

Methylmalonyl-CoA decarboxylase, alpha subunit

75

PTH_1364

mmcI

Desku_1366

Methylmalonyl-CoA decarboxylase, epsilon subunit

82

PTH_1365

mmcJ

Desku_1367

Methylmalonyl-CoA decarboxylase, gamma subunit

56

PTH_1366

mmcK

Desku_1368

Malate dehydrogenase

75

PTH_1367

mmcL

Desku_1369

Transcarboxylase 5S subunit

66

PTH_1368

pykF

Desku_1651

Pyruvate kinase

73

PTH_2214

ppsA

Desku_2615

Pyruvate phosphate dikinase

78

PTH_0903

citE

Desku_2747

Citrate lyase

56

PTH_1335

Corresponding homologs in P. thermopropionicum are included.

Based on 16S rRNA gene sequences, D. kuznetsovii and P. thermopropionicum group in cluster group c and h of the Desulfotomaculum cluster 1, respectively (see Figure 1). P. thermopropionicum is known for its ability to grow with propionate and ethanol in syntrophic association with methanogens. It is not able to grow by sulfate respiration, despite the presence of sulfate reduction genes in the genome [38]. In contrast, D. kuznetsovii is able to grow with propionate (Figure 6) and ethanol with sulfate. However, in the absence of sulfate, it cannot grow in syntrophic association with methanogens. Therefore, differences are expected in genes coding for hydrogenases, formate dehydrogenases, and those involved in sulfate reduction.
Figure 6.

Propionate degradation pathway in D. kuznetsovii based on genomic data. Enzymes are depicted in bold italic. Next to these enzymes are the possible encoding genes, and their locus tags. GCT, Glutaconate CoA-transferase; MCD, Methylmalonyl-CoA decarboxylase; PCC, Propionyl-CoA carboxylase; MCE, Methylmalonyl-CoA epimerase; MCM, Methylmalonyl-CoA mutase; SCS, Succinyl-CoA synthetase; SDH, Succinate dehydrogenase; FHT, Fumarase; MDH, Malate dehydrogenase; OAD, Oxaloacetate decarboxylase; PFL, Pyruvate formate lyase.

Sulfate reduction genes: Figure 7 depicts the sulfate reduction pathway of the two strains. In the genome of D. kuznetsovii two genes (Desku_2103; Desku_3527) are annotated as phosphoadenosine phosphosulfate reductase encoding genes whose corresponding proteins might be involved in assimilatory sulfate metabolism. The P. thermopropionicum genome lacks these genes [39]. Instead, the P. thermopropionicum genome contains an adenylylsulfate kinase gene (PTH_0238). In the dissimilatory sulfate reduction pathway, the two strains both have genes that code for enzymes to reduce sulfate to H2S. However, P. thermopropionicum is missing the gene that codes for an adenylylsulfate reductase beta subunit, which is present in the D. kuznetsovii genome (Desku_1073). Moreover, the gene labeled as a dissimilatory sulfite reductase (dsr) alpha and beta subunit in the P. thermopropionicum genome (PTH_0242) is not similar to dsrA or dsrB from D. kuznetsovii or any other Desulfotomaculum strain. However, it has high similarity to the dsrC gene from D. kuznetsovii, indicating that it is not a dsrA or dsrB gene but a dsrC gene (data not shown). Therefore, the inability of P. thermopropionicum to grow by sulfate respiration is most likely caused by the absence of an adenylylsulfate reductase beta subunit encoding gene and the dsrAB genes.
Figure 7.

Sulfate reduction pathway of D. kuznetsovii and P. thermopropionicum. Depicted in green are genes that code for sulfate reduction enzymes that are present in the genome. Dashed arrows indicate the presence of a subunit encoding gene, but not the presence of all genes required for the enzyme. Dashed dotted arrows are used when no genes were found for the reaction. Abbreviations: APS, adenylylsulfate; DP, diphosphate; PAPS, 3′-Phosphoadenylyl-sulfate (PAPS); redA, reduced acceptor; oxA, oxidized acceptor.

Hydrogenase and formate dehydrogenase genes: Schut and Adams (2009) [40] showed that the trimeric [FeFe]-hydrogenase from Thermotoga maritima oxidizes NADH and ferredoxin simultaneously to produce H2. Similar bifurcating / confurcating [FeFe]-hydrogenases and formate dehydrogenases are present in Syntrophobacter fumaroxidans and P. thermopropionicum [41]. Both generate NADH and ferredoxin during propionate degradation via the methylmalonyl-CoA pathway and might use confurcating hydrogenases and formate dehydrogenases to drive the unfavorable re-oxidation of NADH (E0’=-320mV) by the exergonic re-oxidation of ferredoxin (E0’=-398mV) to produce hydrogen (E0’= -414mV) or formate (E0’= -432mV) that are subsequently transferred to hydrogen and formate scavenging methanogens. Additionally, up-regulation of genes encoding hydrogenases and formate dehydrogenases in P. thermopropionicum was shown during syntrophic growth [42]. The P. thermopropionicum genome contains three [FeFe]-hydrogenases, one [NiFe]-hydrogenase and two formate dehydrogenases. One [FeFe]-hydrogenase (PTH_0668-0670) was shown to be down-regulated during syntrophic growth, while the other two [FeFe]-hydrogenases (PTH_1377-1379 and PTH_2010-2012) were up-regulated. The two formate dehydrogenases of P. thermopropionicum (I, PTH_1711-1714 and II, PTH_2645-2649) were both up-regulated during syntrophic growth [42]. According to TMHMM server v. 2.0 [43] formate dehydrogenase I of P. thermopropionicum has transmembrane helices. Therefore, it might play an essential role in the interspecies transfer of reducing equivalents in syntrophic growth.

The genome of D. kuznetsovii was screened for hydrogenase and formate dehydrogenase encoding gene clusters with BLAST analysis. Pfam search [44] was used to identify motifs in the amino acid sequences and the TMHMM Server v. 2.0 [43] was used to screen for transmembrane helices. The TatP 1.0 Server was used to screen for twin-arginine translocation (Tat) motifs in the N-terminus to predict protein localization in the cell [45]. The incorporation of selenocysteine (SeCys) was examined by RNA loop predictions with Mfold version 3.2 [46, 47]. The predicted RNA loop in the 50–100 bp region downstream of the UGA-codon was compared with the consensus loop described earlier [48].

Compared to P. thermopropionicum, D. kuznetsovii lacks membrane associated formate dehydrogenases and hydrogenases and also lacks [NiFe]-hydrogenase. This might explain why D. kuznetsovii cannot grow in syntrophic relation with methanogens. The genome of D. kuznetsovii indicates the presence of a confurcating selenocysteine-incorporated formate dehydrogenase (Desku_2987-2991), two trimeric confurcating [FeFe]-hydrogenases (Desku_2307-2309, Desku_2995-2997) and two [FeFe]-hydrogenases (Desku_0995, Desku_2934-2935) without NADH-binding sites (Figure 8). Several subunits of these enzymes are related to subunits of NADH dehydrogenase (complex I), including the NADH-binding proteins related to NuoF (Desku_2990, 2308 and 2996) and the electron transfer subunits related to NuoE (Desku_2991, 2935, and 2997) and to NuoG (Desku_2989). In three of the [FeFe]-hydrogenases this NuoG-like domain is fused with the catalytic subunit (Desku_2995, 2307 and 2934). Two of the multimeric hydrogenases are found next to [FeFe]-hydrogenases containing PAS-sensor domains (Desku_2932 and Desku_2994), suggesting they are involved in the regulation of the synthesis of those hydrogenases. All complexes are predicted to be cytoplasmic and not membrane bound.
Figure 8.

Schematic representation of a putative confurcating formate dehydrogenase, two putative confurcating [FeFe]-hydrogenases and two ferredoxin re-oxidizing [FeFe]-hydrogenases in Desulfotomaculum kuznetsovii. Gene locus tag numbers and α-, β-, and γ-subunits are depicted. Moreover, predicted iron-sulfur clusters and metal-binding sites are indicated.

Apart from a possible involvement in the acetate oxidation pathway (Figure 4), it remains unclear for which purpose D. kuznetsovii uses its confurcating formate dehydrogenase and hydrogenases because our genome analysis indicates that pyruvate oxidation during propionate degradation generates formate instead of ferredoxin (Figure 6).

Vitamin synthesis

D. kuznetsovii is able to grow in medium without vitamins [1]. This indicates that D. kuznetsovii is able to synthesize all the vitamins that are required for its metabolism and that vitamin synthesis genes should be present in the genome. Vitamin B12 is essential for the methylmalonyl-CoA pathway and the acetyl-coA pathway. The biosynthesis of cobalamin (vitamin B12) is known to occur from uroporphyrinogen-III to adenosylcobalamin via two possible pathways, the aerobic and anaerobic pathway of the corrinoid ring [49,50]. The D. kuznetsovii genome contains all genes needed for the anaerobic pathway: cysG A (Desku_1520), cysG B (Desku_1460, Desku_1523), cbiA (Desku_1765, Desku_2368), cbiBCDEFGHJLPT (Desku_2369, 1459, 1468, 1467, 1464, 1463, 1462, 1461, 1465, 2370 and 1466, respectively), cobalt reductase (Desku_2757), btuR (Desku_0004, 1209), cobS (Desku_2367) and cobU (Desku_2371). Moreover, D. kuznetsovii has genes to convert glutamyl tRNA to uroporphyrinogen-III, hemABCDL (Desku_1522, 1518, 1521, 1520 and 1522, respectively). The genome also contains some unassigned cobalamin synthesis genes (P47K, Desku_0046, 0053; cbiM, Desku_2905), corrinoid transport proteins (Desku_0693, 702, 2237–2239, 2902–2904, 3025–3027) and, interestingly, two cobN genes (Desku_2189, 2227), genes involved in the aerobic pathway. It is unclear why D. kuznetsovii has these cobN genes, since all anaerobic pathway genes are present in the genome, and it is unclear if the products of these two genes are used for cobalamin synthesis by D. kuznetsovii.

Other vitamin synthesis genes present in the genome of D. kuznetsovii are genes involved in biotin synthesis (vitamin H) (Desku_1295-1297, 2246–2247, 2317), nicotinamide (vitamin B3) synthesis (Desku_0433, 0614, 0662, 0815, 1248, 1417, 1472, 1499, 1925, 1951, 3103, 3121, 3227, 3228, 3231, 3246, 3337), thiamin (vitamin B1) synthesis (0372, 0543, 0545, 2253, 2363, 2639), riboflavin (vitamin B2) synthesis (Desku_1244-1247), and pantothenate (vitamin B5) synthesis (Desku_3262). The genes involved in coenzyme A production from pantothenate are also present in the D. kuznetsovii genome (Desku_1254, 1307, 3145, 3200). Moreover, genes involved in the biosynthesis of pyridoxine (vitamin B6) via the deoxyxylulose 5-phosphate (DXP) independent route were found to be in the genome (Desku_0007, 0008). These genes code for two enzymes that facilitate the conversion of glutamine to the active form of vitamin B6, pyridoxal 5′-phosphate [51].

Menaquinone (vitamin K) and ubiquinone (coenzyme Q10) biosynthesis is important because of the electron transport function in the membranes. The genes that code for the biosynthesis enzymes from polyprenyldiphosphate to menaquinone and ubiquinone are present in the D. kuznetsovii genome (Desku_0124, 0126, 0629, 1551–1554, 1829, 2629 and 3525), except for the genes that code for a 2-polyprenyl-6-methoxyphenol 4-monooxygenase (UbiH) and 2-polyprenyl-3-methyl-6-methoxy-1,4-benzoquinone hydroxylase (UbiF). Additionally, three genes (Desku_1548-1550) could be identified as putative menaquinone biosynthesis genes and are part of a menaquinone biosynthesis gene cluster (Desku_1548-1554). The products of those three genes could be involved in the reactions of the missing UbiH and UbiF encoding genes.

Folate (vitamin B9) biosynthesis is also of great importance for D. kuznetsovii, because it is an essential part of the acetyl-CoA pathway. It is involved in the transfer of one-carbon compounds and can be biosynthesized from chorismate and guanosine triphosphate (GTP) [5255]. Both pathways use a dihydropteroate synthase to produce dihydropteroate. The genome of D. kuznetsovii contains the genes encoding the enzymes involved in the pathway from chorismate to dihydropteroate (Desku_0219, 2268–2269) and from GTP to dihydropteroate (Desku_0210, 0219–0221 and 1419). The gene encoding a phosphatase (Desku_0210) in the D. kuznetsovii genome is probably involved in the removal of phosphate groups from dihydropterine triphosphate as a substitute for an alkaline phosphatase encoding gene, which is not present in the genome. Additionally, the genome contains a bifunctional protein encoding gene (Desku_404) that is expected to be responsible for the production of dihydrofolate (DHF) and the addition of multiple glutamate moieties to DHF or tetrahydrofolate (THF). However, the D. kuznetsovii genome lacks the DHF reductase encoding gene, which is required to reduce DHF to THF. The DHF reductase encoding gene appears to be absent in many microorganisms [56]. Levin et al. (2004) propose that in Halobacterium salinarum a dihydrofolate synthase and a dihydropteroate synthase domain is able to replace the function of the DHF reductase. Additionally, the authors show that when using a BLAST search, homologs of polypeptides can be found in organisms that lack a DHF reductase [56]. However, BLAST results showed no homologous protein encoding gene in the genome of D. kuznetsovii (data not shown). How in D. kuznetsovii DHF is reduced to THF can currently not be deduced from the genome sequence.

Declarations

Acknowledgements

The work conducted by the U.S. Department of Energy Joint Genome Institute is supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC02-05CH11231, and was also supported by grants CW-TOP 700.55.343 and ALW 819.02.014 of the Netherlands Science Foundation (NWO) and grant 323009 of the European Research Council.

Authors’ Affiliations

(1)
Laboratory of Microbiology, Wageningen University
(2)
Department of Aquatic Microbiology, Institute for Biodiversity and Ecosystem Dynamics, University of Amsterdam
(3)
Instituto de Tecnologia Quimica e Biologica, Universidade Nova de Lisboa
(4)
Laboratory of Systems and Synthetic Biology, Wageningen University
(5)
Department of Microbiology, Bremen Institute for Materials Testing
(6)
Winogradsky Institute of Microbiology, Russian Academy of Sciences
(7)
Ecole Polytechnique Federale de Lausanne
(8)
DOE Joint Genome Institute
(9)
Bioscience Division, Los Alamos National Laboratory
(10)
Leibniz Institute, DSMZ - German Collection of Microorganisms and Cell Cultures
(11)
Centre of Biological Engineering, University of Minho

References

  1. Nazina TN, Ivanova AE, Kanchaveli LP, Rozanova EP. A new sporeforming thermophilic methylotrophic sulfate-reducing bacterium, Desulfotomaculum kuznetsovii sp. nov. Mikrobiologiya 1988; 57:823–827.Google Scholar
  2. Stackebrandt E, Sproer C, Rainey FA, Burghardt J, Pauker O, Hippe H. Phylogenetic analysis of the genus Desulfotomaculum: evidence for the mis-classification of Desulfotomaculum guttoideum and description of Desulfotomaculum orientis as Desulfosporosinus orientis gen. nov., comb. nov. Int J Syst Bacteriol 1997; 47:1134–1139. PubMed http://dx.doi.org/10.1099/00207713-47-4-1134View ArticlePubMedGoogle Scholar
  3. Plugge CM, Zhang W, Scholten JC, Stams AJM. Metabolic flexibility of sulfate-reducing bacteria. Front Microbiol 2011; 2:81–87. PubMed http://dx.doi.org/10.3389/fmicb.2011.00081PubMed CentralView ArticlePubMedGoogle Scholar
  4. Field D, Garrity G, Gray T, Morrison N, Selengut J, Sterk P, Tatusova T, Thomson N, Allen MJ, Angiuoli SV, et al. The minimum information about a genome sequence (MIGS) specification. Nat Biotechnol 2008; 26:541–547. PubMed http://dx.doi.org/10.1038/nbt1360PubMed CentralView ArticlePubMedGoogle Scholar
  5. Woese CR, Kandler O, Wheelis ML. Towards a natural system of organisms: proposal for the domains Archaea, Bacteria, and Eucarya. Proc Natl Acad Sci USA 1990; 87:4576–4579. PubMed http://dx.doi.org/10.1073/pnas.87.12.4576PubMed CentralView ArticlePubMedGoogle Scholar
  6. Gibbons NE, Murray RGE. Proposals Concerning the Higher Taxa of Bacteria. Int J Syst Bacteriol 1978; 28:1–6. http://dx.doi.org/10.1099/00207713-28-1-1View ArticleGoogle Scholar
  7. Garrity GM, Holt JG. The Road Map to the Manual. In: Garrity GM, Boone DR, Castenholz RW (eds), Bergey’s Manual of Systematic Bacteriology, Second Edition, Volume 1, Springer, New York, 2001, p. 119–169.View ArticleGoogle Scholar
  8. Murray RGE. The Higher Taxa, or, a Place for Everything…? In: Holt JG (ed), Bergey’s Manual of Systematic Bacteriology, First Edition, Volume 1, The Williams and Wilkins Co., Baltimore, 1984, p. 31–34.Google Scholar
  9. List Editor. List of new names and new combinations previously effectively, but not validly, published. List no. 132. Int J Syst Evol Microbiol 2010; 60:469–472. http://dx.doi.org/10.1099/ijs.0.022855-0
  10. Rainey FA. Class II. Clostridia class nov. In: De Vos P, Garrity G, Jones D, Krieg NR, Ludwig W, Rainey FA, Schleifer KH, Whitman WB (eds), Bergey’s Manual of Systematic Bacteriology, Second Edition, Volume 3, Springer-Verlag, New York, 2009, p. 736.Google Scholar
  11. Skerman VBD, McGowan V, Sneath PHA. Approved Lists of Bacterial Names. Int J Syst Bacteriol 1980; 30:225–420. http://dx.doi.org/10.1099/00207713-30-1-225View ArticleGoogle Scholar
  12. Prévot AR. In: Hauderoy P, Ehringer G, Guillot G, Magrou. J., Prévot AR, Rosset D, Urbain A (eds), Dictionnaire des Bactéries Pathogènes, Second Edition, Masson et Cie, Paris, 1953, p. 1–692.Google Scholar
  13. Rogosa M. Peptococcaceae, a new family to include the Gram-positive, anaerobic cocci of the genera Peptococcus, Peptostreptococcus and Ruminococcus. Int J Syst Bacteriol 1971; 21:234–237. http://dx.doi.org/10.1099/00207713-21-3-234View ArticleGoogle Scholar
  14. Campbell LL, Postgate JR. Classification of the spore-forming sulfate-reducing bacteria. Bacteriol Rev 1965; 29:359–363. PubMedPubMed CentralPubMedGoogle Scholar
  15. Campbell LL. Genus IV. Desulfotomaculum Campbell and Postgate 1965, 361. In: Buchanan RE, Gibbons NE (eds), Bergey’s Manual of Determinative Bacteriology, Eighth Edition, The Williams and Wilkins Co., Baltimore, 1974, p. 572–573.Google Scholar
  16. Validation of the publication of new names and new combinations previously effectively published outside the IJSB. List no. 35. Int J Syst Bacteriol 1990; 40:470–471. http://dx.doi.org/10.1099/00207713-40-4-470
  17. Isaksen MF, Bak F, Jorgensen BB. Thermophilic sulfate-reducing bacteria in cold marine sediment. Microb Ecol 1994; 14:1–8. http://dx.doi.org/10.1111/j.1574-6941.1994.tb00084.xView ArticleGoogle Scholar
  18. Goorissen HP, Boschker HT, Stams AJM, Hansen TA. Isolation of thermophilic Desulfotomaculum strains with methanol and sulfite from solfataric mud pools, and characterization of Desulfotomaculum solfataricum sp. nov. Int J Syst Evol Microbiol 2003; 53:1223–1229. PubMed http://dx.doi.org/10.1099/ijs.0.02476-0View ArticlePubMedGoogle Scholar
  19. Ashburner M, Ball CA, Blake JA, Botstein D, Butler H, Cherry JM, Davis AP, Dolinski K, Dwight SS, Eppig JT, et al. Gene ontology: tool for the unification of biology. The Gene Ontology Consortium. Nat Genet 2000; 25:25–29. PubMed http://dx.doi.org/10.1038/75556PubMed CentralView ArticlePubMedGoogle Scholar
  20. Pagani I, Liolios K, Jansson J, Chen IM, Smirnova T, Nosrat B, Markowitz VM, Kyrpides NC. The Genomes OnLine Database (GOLD) v.4: status of genomic and metagenomic projects and their associated metadata. Nucleic Acids Res 2012; 40(Database issue):D571–D579. PubMed http://dx.doi.org/10.1093/nar/gkr1100PubMed CentralView ArticlePubMedGoogle Scholar
  21. JGI website. http://www.jgi.doe.gov.
  22. The Phred/Phrap/Consed software package. http://www.phrap.com.
  23. Zerbino DR, Birney E. Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Genome Res 2008; 18:821–829. PubMed http://dx.doi.org/10.1101/gr.074492.107PubMed CentralView ArticlePubMedGoogle Scholar
  24. Han C, Chain P. Finishing repeat regions automatically with Dupfinisher. In: H.R. A, H. V, editors2006 June 26–29, 2006. CSREA Press. p 141–6.
  25. Lapidus A, LaButti K, Foster B, Lowry S, Trong S, Goltsman E. POLISHER: An effective tool for using ultra short reads in microbial genome assembly and finishing. Marco Island, FL: AGBT; 2008.Google Scholar
  26. Hyatt D, Chen GL, Locascio PF, Land ML, Larimer FW, Hauser LJ. Prodigal: prokaryotic gene recognition and translation initiation site identification. BMC Bioinformatics 2010; 11:119. PubMed http://dx.doi.org/10.1186/1471-2105-11-119PubMed CentralView ArticlePubMedGoogle Scholar
  27. Pati A, Ivanova NN, Mikhailova N, Ovchinnikova G, Hooper SD, Lykidis A, Kyrpides NC. GenePRIMP: a gene prediction improvement pipeline for prokaryotic genomes. Nat Methods 2010; 7:455–457. PubMed http://dx.doi.org/10.1038/nmeth.1457View ArticlePubMedGoogle Scholar
  28. Markowitz VM, Mavromatis K, Ivanova NN, Chen IM, Chu K, Kyrpides NC. IMG ER: a system for microbial genome annotation expert review and curation. Bioinformatics 2009; 25:2271–2278. PubMed http://dx.doi.org/10.1093/bioinformatics/btp393View ArticlePubMedGoogle Scholar
  29. Tourova TP, Kuznetsov BB, Novikova EV, Poltaraus AB, Nazina TN. Heterogeneity of the nucleotide sequences of the 16 S rRNA genes of the type strain of Desulfotomaculum kuznetsovii. Mikrobiologiya 2001; 70:788–795.Google Scholar
  30. Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 1997; 25:3389–3402. PubMed http://dx.doi.org/10.1093/nar/25.17.3389PubMed CentralView ArticlePubMedGoogle Scholar
  31. Altschul SF, Wootton JC, Gertz EM, Agarwala R, Morgulis A, Schaffer AA, Yu YK. Protein database searches using compositionally adjusted substitution matrices. FEBS J 2005; 272:5101–5109. PubMed http://dx.doi.org/10.1111/j.1742-4658.2005.04945.xPubMed CentralView ArticlePubMedGoogle Scholar
  32. Goevert D, Conrad R. Carbon isotope fractionation by sulfate-reducing bacteria using different pathways for the oxidation of acetate. Environ Sci Technol 2008; 42:7813–7817. PubMed http://dx.doi.org/10.1021/es800308zView ArticlePubMedGoogle Scholar
  33. Spormann AM, Thauer RK. Anaerobic acetate oxidation to CO2 in Desulfotomaculum acetoxidans. Arch Microbiol 1988; 150:374–380. http://dx.doi.org/10.1007/BF00408310View ArticleGoogle Scholar
  34. Spring S, Lapidus A, Schroder M, Gleim D, Sims D, Meincke L, Glavina Del Rio T, Tice H, Copeland A, Cheng JF, et al. Complete genome sequence of Desulfotomaculum acetoxidans type strain (5575). Stand Genomic Sci 2009; 1:242–253. PubMed http://dx.doi.org/10.4056/sigs.39508PubMed CentralView ArticlePubMedGoogle Scholar
  35. Goorissen HP, Stams AJM, Hansen TA. Methanol dissimilation in Desulfotomaculum kuznetsovii PhD dissertation: Thermophilic methanol utilization by sulfate reducing bacteria 2002;Chapter 3:55–61.
  36. Pierce E, Xie G, Barabote RD, Saunders E, Han CS, Detter JC, Richardson P, Brettin TS, Das A, Ljungdahl LG, et al. The complete genome sequence of Moorella thermoacetica (f. Clostridium thermoaceticum). Environ Microbiol 2008; 10:2550–2573. PubMed http://dx.doi.org/10.1111/j.1462-2920.2008.01679.xPubMed CentralView ArticlePubMedGoogle Scholar
  37. van der Meijden P, Heythuysen HJ, Pouwels A, Houwen F, van der Drift C, Vogels GD. Methyltransferases involved in methanol conversion by Methanosarcina barkeri. Arch Microbiol 1983; 134:238–242. PubMed http://dx.doi.org/10.1007/BF00407765View ArticlePubMedGoogle Scholar
  38. Imachi H, Sekiguchi Y, Kamagata Y, Hanada S, Ohashi A, Harada H. Pelotomaculum thermopropionicum gen. nov., sp. nov., an anaerobic, thermophilic, syntrophic propionate-oxidizing bacterium. Int J Syst Evol Microbiol 2002; 52:1729–1735. PubMed http://dx.doi.org/10.1099/ijs.0.02212-0PubMedGoogle Scholar
  39. Kosaka T, Kato S, Shimoyama T, Ishii S, Abe T, Watanabe K. The genome of Pelotomaculum thermopropionicum reveals niche-associated evolution in anaerobic microbiota. Genome Res 2008; 18:442–448. PubMed http://dx.doi.org/10.1101/gr.7136508PubMed CentralView ArticlePubMedGoogle Scholar
  40. Schut GJ, Adams MW. The iron-hydrogenase of Thermotoga maritima utilizes ferredoxin and NADH synergistically: a new perspective on anaerobic hydrogen production. J Bacteriol 2009; 191:4451–4457. PubMed http://dx.doi.org/10.1128/JB.01582-08PubMed CentralView ArticlePubMedGoogle Scholar
  41. Müller N, Worm P, Schink B, Stams AJM, Plugge CM. Syntrophic butyrate and propionate oxidation processes: from genomes to reaction mechanisms. Environmental Microbiology Reports 2010; 2:489–499. http://dx.doi.org/10.1111/j.1758-2229.2010.00147.xView ArticlePubMedGoogle Scholar
  42. Kato S, Kosaka T, Watanabe K. Substrate-dependent transcriptomic shifts in Pelotomaculum thermopropionicum grown in syntrophic co-culture with Methanothermobacter thermautotrophicus. Microb Biotechnol 2009; 2:575–584. PubMed http://dx.doi.org/10.1111/j.1751-7915.2009.00102.xPubMed CentralView ArticlePubMedGoogle Scholar
  43. DTU. 2009 Center for biological sequence analysis: Technical University of Denmark. http://www.cbs.dtu.dk/services/TMHMM
  44. Sanger Institute CU. 2009. http://pfam.sanger.ac.uk/search.
  45. Bendtsen JD, Nielsen H, Widdick D, Palmer T, Brunak S. Prediction of twin-arginine signal peptides. BMC Bioinformatics 2005; 6:167. PubMed http://dx.doi.org/10.1186/1471-2105-6-167PubMed CentralView ArticlePubMedGoogle Scholar
  46. Mathews DH, Sabina J, Zuker M, Turner DH. Expanded sequence dependence of thermodynamic parameters improves prediction of RNA secondary structure. J Mol Biol 1999; 288:911–940. PubMed http://dx.doi.org/10.1006/jmbi.1999.2700View ArticlePubMedGoogle Scholar
  47. Zuker M. Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res 2003; 31:3406–3415. PubMed http://dx.doi.org/10.1093/nar/gkg595PubMed CentralView ArticlePubMedGoogle Scholar
  48. Zhang Y, Gladyshev VN. An algorithm for identification of bacterial selenocysteine insertion sequence elements and selenoprotein genes. Bioinformatics 2005; 21:2580–2589. PubMed http://dx.doi.org/10.1093/bioinformatics/bti400View ArticlePubMedGoogle Scholar
  49. Roth JR, Lawrence JG, Bobik TA. Cobalamin (co-enzyme B12): synthesis and biological significance. Annu Rev Microbiol 1996; 50:137–181. PubMed http://dx.doi.org/10.1146/annurev.micro.50.1.137View ArticlePubMedGoogle Scholar
  50. Roessner CA, Santander PJ, Scott AI. Multiple biosynthetic pathways for vitamin B12: variations on a central theme. Vitam Horm 2001; 61:267–297. PubMed http://dx.doi.org/10.1016/S0083-6729(01)61009-4View ArticlePubMedGoogle Scholar
  51. Fitzpatrick TB, Amrhein N, Kappes B, Macheroux P, Tews I, Raschle T. Two independent routes of de novo vitamin B6 biosynthesis: not that different after all. Biochem J 2007; 407:1–13. PubMed http://dx.doi.org/10.1042/BJ20070765View ArticlePubMedGoogle Scholar
  52. Bermingham A, Derrick JP. The folic acid biosynthesis pathway in bacteria: evaluation of potential for antibacterial drug discovery. Bioessays 2002; 24:637–648. PubMed http://dx.doi.org/10.1002/bies.10114View ArticlePubMedGoogle Scholar
  53. Dosselaere F, Vanderleyden J. A metabolic node in action: chorismate-utilizing enzymes in micro-organisms. Crit Rev Microbiol 2001; 27:75–131. PubMed http://dx.doi.org/10.1080/20014091096710View ArticlePubMedGoogle Scholar
  54. Hu SI, Drake HL, Wood HG. Synthesis of acetyl coenzyme A from carbon monoxide, methyltetrahydrofolate, and coenzyme A by enzymes from Clostridium thermoaceticum. J Bacteriol 1982; 149:440–448. PubMedPubMed CentralPubMedGoogle Scholar
  55. Stupperich E, Konle R. Corrinoid-Dependent Methyl Transfer Reactions Are Involved in Methanol and 3,4-Dimethoxybenzoate Metabolism by Sporomusa ovata. Appl Environ Microbiol 1993; 59:3110–3116. PubMedPubMed CentralPubMedGoogle Scholar
  56. Levin I, Giladi M, Altman-Price N, Ortenberg R, Mevarech M. An alternative pathway for reduced folate biosynthesis in bacteria and halophilic archaea. Mol Microbiol 2004; 54:1307–1318. PubMed http://dx.doi.org/10.1111/j.1365-2958.2004.04339.xView ArticlePubMedGoogle Scholar

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