Cloning of Genes Coding for the Three Subunits of Thiocyanate Hydrolase of Thiobacillus thioparus THI 115 and Their Evolutionary Relationships to Nitrile Hydratase

Abstract

Thiocyanate hydrolase is a newly found enzyme fromThiobacillus thioparus THI 115 that converts thiocyanate to carbonyl sulfide and ammonia (Y. Katayama, Y. Narahara, Y. Inoue, F. Amano, T. Kanagawa, and H. Kuraishi, J. Biol. Chem. 267:9170–9175, 1992). We have cloned and sequenced the scngenes that encode the three subunits of the enzyme. ThescnB, scnA, and scnC genes, arrayed in this order, contained open reading frames encoding sequences of 157, 126, and 243 amino acid residues, respectively, for the β, α, and γ subunits, respectively. Each open reading frame was preceded by a typical Shine-Dalgarno sequence. The deduced amino-terminal peptide sequences for the three subunits were in fair agreement with the chemically determined sequences. The protein molecular mass calculated for each subunit was compatible with that determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. From a computer analysis, thiocyanate hydrolase showed significant homologies to bacterial nitrile hydratases known to convert nitrile to the corresponding amide, which is further hydrolyzed by amidase to form acid and ammonia. The two enzymes were homologous over regions corresponding to almost the entire coding regions of the genes: the β and α subunits of thiocyanate hydrolase were homologous to the amino- and carboxyl-terminal halves of the β subunit of nitrile hydratase, and the γ subunit of thiocyanate hydrolase was homologous to the α subunit of nitrile hydratase. Comparisons of the catalytic properties of the two homologous enzymes support the model for the reaction steps of thiocyanate hydrolase that was previously presented on the basis of biochemical analyses.

New insights into the genetic and metabolic diversity of thiocyanate-degrading microbial consortia

“…The specific enzyme that mediates this reaction, SCNhydrolase (SCNase), first identified in the chemolithotrophic bacterium T. thioparus THI 115 (Katayama et al 1992), exhibits significant homology to bacterial nitrile hydratases (Katayama et al 1998). The latter study was able to clone and sequence the genes encoding the β, ɑ and γ subunits of this enzyme; the scnB, scnA and scnC genes respectively.…”

Bioreactor microbial ecosystems for thiocyanate and cyanide degradation unravelled with genome‐resolved metagenomics

“…Activated Sludge Tailings Effluent Removal: ASTER TM process, Biomin Limited; Consort Mine in Barberton, South Africa; Suzdal Mine, Kazakhstan; and Homestake Mine, SD, USA) (van Buuren et al, 2011). The microbiological and molecular study of SCN − and CN − degradation has focused on isolates lacking sequenced genomes (Katayama et al, 1998;Wood et al, 1998;du Plessis et al, 2001;Sorokin et al, 2004;Arakawa et al, 2007;Hussain et al, 2013). Molecular fingerprinting of SCN − degrading consortia (Quan et al, 2006;Felföldi et al, 2010;Huddy et al, 2015) provides the only knowledge at the community level to date.…”

Exchange of carbonyl sulfide (OCS) between soils and atmosphere under various CO2 concentrations

“…Of special interest is the light-dependent and obviously abiotic OCS production reported by Whelan and Rhew [2015]. Two biotic sources have been reported, production from thiocyanate by thiocyanate hydrolase [Katayama et al, 1992;Katayama et al, 1998] and production from carbon disulfide by carbon disulfide hydrolase [Smith and Kelly, 1988;Smeulders et al, 2011]. Lehmann and Conrad [1996] also observed an increase of OCS emission from soils after the addition of thiocyanate.…”