The genome sequence of the obligately chemolithoautotrophic, facultatively anaerobic bacterium Thiobacillus denitrificans

Abstract

The complete genome sequence of Thiobacillus denitrificans ATCC 25259 is the first to become available for an obligately chemolithoautotrophic, sulfur-compound-oxidizing, beta-proteobacterium. Analysis of the 2,909,809-bp genome will facilitate our molecular and biochemical understanding of the unusual metabolic repertoire of this bacterium, including its ability to couple denitrification to sulfur-compound oxidation, to catalyze anaerobic, nitrate-dependent oxidation of Fe(II) and U(IV), and to oxidize mineral electron donors. Notable genomic features include (i) genes encoding c-type cytochromes totaling 1 to 2 percent of the genome, which is a proportion greater than for almost all bacterial and archaeal species sequenced to date, (ii) genes encoding two [NiFe]hydrogenases, which is particularly significant because no information on hydrogenases has previously been reported for T. denitrificans and hydrogen oxidation appears to be critical for anaerobic U(IV) oxidation by this species, (iii) a diverse complement of more than 50 genes associated with sulfur-compound oxidation (including sox genes, dsr genes, and genes associated with the AMP-dependent oxidation of sulfite to sulfate), some of which occur in multiple (up to eight) copies, (iv) a relatively large number of genes associated with inorganic ion transport and heavy metal resistance, and (v) a paucity of genes encoding organic-compound transporters, commensurate with obligate chemolithoautotrophy. Ultimately, the genome sequence of T. denitrificans will enable elucidation of the mechanisms of aerobic and anaerobic sulfur-compound oxidation by beta-proteobacteria and will help reveal the molecular basis of this organism's role in major biogeochemical cycles (i.e., those involving sulfur, nitrogen, and carbon) and groundwater restoration.

FIG. 1.

Schematic circular diagram of the T. denitrificans ATCC 25259 genome. Outer circle, predicted coding regions on the forward strand, color-coded by role categories (dark gray, hypothetical proteins; light gray, conserved hypothetical and unknown function; brown, general function; red, DNA replication and repair; green, energy metabolism; blue, carbon and carbohydrate metabolism; cyan, lipid metabolism; magenta, transcription; yellow, translation; orange, amino acid metabolism; pink, metabolism of cofactors and vitamins; light red, purine and pyrimidine metabolism; lavender, signal transduction; sky blue, cellular processes; pale green, structural RNAs); second circle, predicted coding regions on the reverse strand, color coded as for the outer circle; third and fourth circles, coding regions (on forward and reverse strands) predicted to be involved in denitrification (blue), sulfur-compound oxidation (red), hydrogen oxidation (green), autotrophic carbon assimilation (orange), and metal ion transport-resistance (brown); fifth and sixth circles, coding regions found to have a CXXCH heme-binding motif and therefore potentially encoding c-type cytochromes; seventh circle, deviation from the average G+C; eighth circle, GC skew (olive, positive; purple, negative).

FIG. 2.

Schematic overview of key genes/enzymes putatively associated with sulfur-compound oxidation in T. denitrificans. Genes in parenthesis have been shown to be lesser expressed paralogs (this study). The biochemical roles of a number of gene products represented in this figure have not been experimentally demonstrated in T. denitrificans and are uncertain. Sulfide:quinone oxidoreductase is not proposed to catalyze the direct oxidation of sulfide to sulfite but rather may participate in an indirect pathway (20). The arrow between thiosulfate and sulfate (right side) represents the possibility that SoxB catalyzes a sulfate thiohydrolase reaction (28) in T. denitrificans. APAT, APS:phosphate adenylyltransferase.

FIG. 3.

Phylogenetic relationships among the eight putative DsrC proteins encoded in T. denitrificans ATCC 25259 and the top BLASTP matches from the GenBank nr database for Tbd2480. Of the proteins represented in this figure that are not from T. denitrificans, more than 70% are known or predicted to be DsrC or more broadly related to sulfite reductases (indicated in boldface type). For limbs that show species names rather than GenBank accession numbers, the corresponding accession numbers are as follows: A. vinosum (AAC35399.1), M. magnetotacticum (ZP 00052645.1), Magnetococcus sp. (ZP 00287929.1), C. tepidum (NP 663123.1), T. norvegica (CAC36215.1), D. desulfuricans (ZP 00130056.2), and D. vulgaris (YP 011988).

FIG.4.

Phylogenetic relationships among predicted amino acid sequences for HynS (A), Isp1 (B), Isp2 (C), and HynL (D) in T. denitrificans and the best BLASTP matches from the GenBank nr database. For limbs that show species names rather than GenBank accession numbers, the corresponding accession numbers are as follows: (HynS) A. vinosum (AAU93828.1), T. roseopersicina (AAC38281.1), A. aeolicus (NP 213658.1); (Isp1) A. vinosum (AAU93829.2), T. roseopersicina (AAC38283.1), A. aeolicus (NP 213657.1), D. vulgaris Hmc5 (YP 009755), A. ambivalens (CAC86885.1), D. desulfuricans NarI (ZP 00128546.1); (Isp2) A. vinosum (AAY89333.1), T. roseopersicina (AAC38284.1), A. aeolicus (NP213656.1), A. ambivalens (CAC86886.1), Polaromonas sp. (ZP 00503323.1), C. aurantiacus (ZP 00356812), A. vinosum DsrK (AAC35401.2), C. tepidum DsrK (NP 663117.1), D. vulgaris Hmc6 (YP 009754.1); (HynL) A. vinosum (AAY89334.1), T. roseopersicina (AAC38282.1), A. aeolicus (NP213655.1).