Regulation of a Novel
            <i>Acidithiobacillus caldus</i>
            Gene Cluster Involved in Metabolism of Reduced Inorganic Sulfur Compounds

Acidithiobacillus caldus is a moderately thermophilic, acidophilic bacterium that has been reported to be the dominant sulfur oxidizer in stirred-tank processes used to treat gold-bearing arsenopyrite ores. It is also widely distributed in heap reactors used for the extraction of metals from ores. Not only are these bacteria commercially important, they have an interesting physiology, the study of which has been restricted by the nonavailability of defined mutants. A recently reported conjugation system based on the broad-host-range IncW plasmids pSa and R388 was used to transfer mobilizable narrow-host-range suicide plasmid vectors containing inactivated and partially deleted chromosomal genes from Escherichia coli to A. caldus. Through the dual use of a selectable kanamycin resistance gene and a hybridization probe made from a deleted portion of the target chromosomal gene, single-and double-recombinant mutants of A. caldus were isolated. The functionality of the gene inactivation system was shown by the construction of A. caldus arsB and tetH mutants, and the effects of these mutations on cell growth in the presence of arsenic and by means of tetrathionate oxidation were demonstrated.

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“…These are the genes conferring arsenic resistance (15) and the genes for tetrathionate utilization (26).…”

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confidence: 99%

Construction of arsB and tetH Mutants of the Sulfur-Oxidizing Bacterium Acidithiobacillus caldus by Marker Exchange

Zyl

Munster

Rawlings

2008

Appl Environ Microbiol

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“…ambivalens. 23,25,27) Fig. 1 shows the alignment of the primary structure of the three 4THases excluding their putative signal peptides.…”

Section: Resultsmentioning

confidence: 99%

The sole cysteine residue (Cys301) of tetrathionate hydrolase from Acidithiobacillus ferrooxidans does not play a role in enzyme activity

Kanao

Nakayama

Kato

et al. 2014

Bioscience, Biotechnology, and Biochemistry

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Cysteine residues are absolutely indispensable for the reactions of almost all enzymes involved in the dissimilatory oxidation pathways of reduced inorganic sulfur compounds. Tetrathionate hydrolase from the acidophilic iron- and sulfur-oxidizing bacterium Acidithiobacillus ferrooxidans (Af-Tth) catalyzes tetrathionate hydrolysis to generate elemental sulfur, thiosulfate, and sulfate. Af-Tth is a key enzyme in the dissimilatory sulfur oxidation pathway in this bacterium. Only one cysteine residue (Cys301) has been identified in the deduced amino acid sequence of the Af-Tth gene. In order to clarify the role of the sole cysteine residue, a site-specific mutant enzyme (C301A) was generated. No difference was observed in the retention volumes of the wild-type and mutant Af-Tth enzymes by gel-filtration column chromatography, and surprisingly the enzyme activities measured in the cysteine-deficient and wild-type enzymes were the same. These results suggest that the sole cysteine residue (Cys301) in Af-Tth is involved in neither the tetrathionate hydrolysis reaction nor the subunit assembly. Af-Tth may thus have a novel cysteine-independent reaction mechanism.

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“…Tetrathionate can be hydrolyzed to thiosulfate, sulfate, and sulfur by tetrathionate hydrolase (TetH) (7)(8)(9). Compared with TetH from the sulfur-oxidizing strain Acidithiobacillus caldus (10,11), TetH from A. ferrooxidans was speculated to be phylogenetically distinct (11,12) and even diverse in its catalytic reaction on account of different cofactor requirements (10,13). In addition, the subcellular location of TetH in A. ferrooxidans has not been certain until now because it has been reported as a periplasmic (8), outer membrane-associated (12,14), and extracellular (15) protein.…”

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Construction and Characterization of tetH Overexpression and Knockout Strains of Acidithiobacillus ferrooxidans

Liu

Wang

et al. 2014

J Bacteriol

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Acidithiobacillus ferrooxidans is a major participant in consortia of microorganisms used for bioleaching. It can obtain energy from the oxidation of Fe 2؉ , H 2 , S 0 , and various reduced inorganic sulfur compounds (RISCs). Tetrathionate is a key intermediate during RISC oxidation, hydrolyzed by tetrathionate hydrolase (TetH), and used as sole energy source. In this study, a tetH knockout (⌬tetH) mutant and a tetH overexpression strain were constructed and characterized. The tetH overexpression strain grew better on sulfur and tetrathionate and possessed a higher rate of tetrathionate utilization and TetH activity than the wild type. However, its cell yields on tetrathionate were much lower than those on sulfur. The ⌬tetH mutant could not grow on tetrathionate but could proliferate on sulfur with a lower cell yield than the wild type's, which indicated that tetrathionate hydrolysis is mediated only by TetH, encoded by tetH. The ⌬tetH mutant could survive in ferrous medium with an Fe 2؉ oxidation rate similar to that of the wild type. For the tetH overexpression strain, the rate was relatively higher than that of the wild type. The reverse transcription-quantitative PCR (qRT-PCR) results showed that tetH and doxD2 acted synergistically, and doxD2 was considered important in thiosulfate metabolism. Of the two sqr genes, AFE_0267 seemed to play as important a role in sulfide oxidation as AFE_1792. This study not only provides a substantial basis for studying the function of the tetH gene but also may serve as a model to clarify other candidate genes involved in sulfur oxidation in this organism.

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Regulation of a Novel Acidithiobacillus caldus Gene Cluster Involved in Metabolism of Reduced Inorganic Sulfur Compounds

Cited by 60 publications

References 41 publications

Construction of arsB and tetH Mutants of the Sulfur-Oxidizing Bacterium Acidithiobacillus caldus by Marker Exchange

Construction of arsB and tetH Mutants of the Sulfur-Oxidizing Bacterium Acidithiobacillus caldus by Marker Exchange

The sole cysteine residue (Cys301) of tetrathionate hydrolase from Acidithiobacillus ferrooxidans does not play a role in enzyme activity

Construction and Characterization of tetH Overexpression and Knockout Strains of Acidithiobacillus ferrooxidans

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