1887

Abstract

Strain THI201, a member of the alphaproteobacteria, is a novel thiocyanate (SCN)-degrading bacterium isolated from lake water enriched with potassium thiocyanate (KSCN). This bacterium carries the enzyme thiocyanate hydrolase (SCNase) that hydrolyses thiocyanate to carbonyl sulfide and ammonia. Characterization of both native and recombinant SCNase revealed properties different from known SCNases regarding subunit structure and thermostability: SCNase of strain THI201 was composed of a single protein and thermostable. We cloned and sequenced the corresponding gene and determined a protein of 457 amino acids of molecular mass 50 267 Da. Presence of a twin-arginine (Tat) signal sequence of 32 amino acids was found upstream of SCNase. The deduced amino acid sequence of SCNase showed 83 % identity to that of a putative uncharacterized protein of ATCC 25259, but no significant identity to those of three subunits of SCNase from strain THI115. The specific activities of native and recombinant enzyme were 0.32 and 4–15 µmol min (mg protein), respectively. The maximum activity of SCNase was found in the temperature range 30–70 °C. The thiocyanate-hydrolysing activity in both enzymes was decreased by freeze–thawing, although 25–100 % of the activity of recombinant protein could be retrieved by treating the enzyme at 60 °C for 15 min. Furthermore, both native and recombinant enzymes retained the activity after pre-treatment of the protein solution at temperatures up to 70 °C.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.063339-0
2013-11-01
2020-01-19
Loading full text...

Full text loading...

/deliver/fulltext/micro/159/11/2294.html?itemId=/content/journal/micro/10.1099/mic.0.063339-0&mimeType=html&fmt=ahah

References

  1. Arakawa T., Kawano Y., Kataoka S., Katayama Y., Kamiya N., Yohda M., Odaka M..( 2007;). Structure of thiocyanate hydrolase: a new nitrile hydratase family protein with a novel five-coordinate cobalt(III) center. J Mol Biol366:1497–1509 [CrossRef][PubMed]
    [Google Scholar]
  2. Bezsudnova E. Y., Sorokin D. Y., Tikhonova T. V., Popov V. O..( 2007;). Thiocyanate hydrolase, the primary enzyme initiating thiocyanate degradation in the novel obligately chemolithoautotrophic halophilic sulfur-oxidizing bacterium Thiohalophilus thiocyanoxidans. Biochim Biophys Acta1774:1563–1570 [CrossRef][PubMed]
    [Google Scholar]
  3. Burow M., Bergner A., Gershenzon J., Wittstock U..( 2007;). Glucosinolate hydrolysis in Lepidium sativum – identification of the thiocyanate-forming protein. Plant Mol Biol63:49–61 [CrossRef][PubMed]
    [Google Scholar]
  4. Chollet R., Anderson L. L..( 1977;). Conformational changes associated with the reversible cold inactivation of ribulose-1,5-bisphosphate carboxylase-oxygenase. Biochim Biophys Acta482:228–240 [CrossRef][PubMed]
    [Google Scholar]
  5. Erez T., Gdalevsky G. Ya., Torchinsky Y. M., Phillips R. S., Parola A. H..( 1998;). Cold inactivation and dissociation into dimers of Escherichia coli tryptophanase and its W330F mutant form. Biochim Biophys Acta1384:365–372 [CrossRef][PubMed]
    [Google Scholar]
  6. Happold F. C., Jones G. L., Pratt D. B..( 1958;). Utilization of thiocyanate by Thiobacillus thioparus and T. thiocyanoxidans. Nature182:266–267 [CrossRef][PubMed]
    [Google Scholar]
  7. Jürgen B., Lin H. Y., Riemschneider S., Scharf C., Neubauer P., Schmid R., Hecker M., Schweder T..( 2000;). Monitoring of genes that respond to overproduction of an insoluble recombinant protein in Escherichia coli glucose-limited fed-batch fermentations. Biotechnol Bioeng70:217–224 [CrossRef][PubMed]
    [Google Scholar]
  8. Kataoka S., Arakawa T., Hori S., Katayama Y., Hara Y., Matsushita Y., Nakayama H., Yohda M., Nyunoya H. et al.( 2006;). Functional expression of thiocyanate hydrolase is promoted by its activator protein, P15K. FEBS Lett580:4667–4672 [CrossRef][PubMed]
    [Google Scholar]
  9. Katayama Y., Kuraishi H..( 1978;). Characteristics of Thiobacillus thioparus and its thiocyanate assimilation. Can J Microbiol24:804–810 [CrossRef][PubMed]
    [Google Scholar]
  10. Katayama Y., Narahara Y., Inoue Y., Amano F., Kanagawa T., Kuraishi H..( 1992;). A thiocyanate hydrolase of Thiobacillus thioparus. A novel enzyme catalyzing the formation of carbonyl sulfide from thiocyanate. J Biol Chem267:9170–9175[PubMed]
    [Google Scholar]
  11. Katayama Y., Hiraishi A., Kuraishi H..( 1995;). Paracoccus thiocyanatus sp. nov., a new species of thiocyanate-utilizing facultative chemolithotroph, and transfer of Thiobacillus versutus to the genus Paracoccus as Paracoccus versutus comb. nov. with emendation of the genus. Microbiology141:1469–1477 [CrossRef][PubMed]
    [Google Scholar]
  12. Katayama Y., Matsushita Y., Kaneko M., Kondo M., Mizuno T., Nyunoya H..( 1998;). Cloning of genes coding for the three subunits of thiocyanate hydrolase of Thiobacillus thioparus THI 115 and their evolutionary relationships to nitrile hydratase. J Bacteriol180:2583–2589[PubMed]
    [Google Scholar]
  13. Katayama Y., Hashimoto K., Nakayama H., Mino H., Nojiri M., Ono T. A., Nyunoya H., Yohda M., Takio K., Odaka M..( 2006;). Thiocyanate hydrolase is a cobalt-containing metalloenzyme with a cysteine-sulfinic acid ligand. J Am Chem Soc128:728–729 [CrossRef][PubMed]
    [Google Scholar]
  14. Kelly D. P., Malin G., Wood A. P..( 1993;). Microbial transformations and biogeochemical cycling of one-carbon substrates containing sulphur, nitrogen or halogens. Microbial Growth on C1 Compounds47–63 Murrell J. C., Kelly D. P.. Andover: Intercept;
    [Google Scholar]
  15. Laemmli U. K..( 1970;). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature227:680–685 [CrossRef][PubMed]
    [Google Scholar]
  16. Lowry O. H., Rosebrough N. J., Farr A. L., Randall R. J..( 1951;). Protein measurement with the Folin phenol reagent. J Biol Chem193:265–275[PubMed]
    [Google Scholar]
  17. Meulenberg R., Pronk J. T., Frank J., Hazeu W., Bos P., Kuenen J. G..( 1992;). Purification and partial characterization of a thermostable trithionate hydrolase from the acidophilic sulphur oxidizer Thiobacillus acidophilus. Eur J Biochem209:367–374 [CrossRef][PubMed]
    [Google Scholar]
  18. Miyanaga A., Fushinobu S., Ito K., Wakagi T..( 2001;). Crystal structure of cobalt-containing nitrile hydratase. Biochem Biophys Res Commun288:1169–1174 [CrossRef][PubMed]
    [Google Scholar]
  19. Nagashima S., Nakasako M., Dohmae N., Tsujimura M., Takio K., Odaka M., Yohda M., Kamiya N., Endo I..( 1998;). Novel non-heme iron center of nitrile hydratase with a claw setting of oxygen atoms. Nat Struct Biol5:347–351 [CrossRef][PubMed]
    [Google Scholar]
  20. Rinas U..( 1996;). Synthesis rates of cellular proteins involved in translation and protein folding are strongly altered in response to overproduction of basic fibroblast growth factor by recombinant Escherichia coli. Biotechnol Prog12:196–200 [CrossRef][PubMed]
    [Google Scholar]
  21. Saito H., Miura K. I..( 1963;). Preparation of transforming deoxyribonucleic acid by phenol treatment. Biochim Biophys Acta72:619–629 [CrossRef][PubMed]
    [Google Scholar]
  22. Saito A., Mitsui H., Hattori R., Minamisawa K., Hattori T..( 1998;). Slow-growing and oligotrophic soil bacteria phylogetically close to Bradyrhizobiumjaponicum. FEMS Microbiol Ecol25:277–286 [CrossRef]
    [Google Scholar]
  23. Sareen D., Sharma R., Vohra R. M..( 2001;). Chaperone-assisted overexpression of an active D-carbamoylase from Agrobacterium tumefaciens AM 10. Protein Expr Purif23:374–379 [CrossRef][PubMed]
    [Google Scholar]
  24. Sörbo B..( 1987;). Sulfate: turbidimetric and nephelometric methods. Methods Enzymol143:3–6 [CrossRef][PubMed]
    [Google Scholar]
  25. Stevens J. M., Rao Saroja N., Jaouen M., Belghazi M., Schmitter J. M., Mansuy D., Artaud I., Sari M. A..( 2003;). Chaperone-assisted expression, purification, and characterization of recombinant nitrile hydratase NI1 from Comamonas testosteroni. Protein Expr Purif29:70–76 [CrossRef][PubMed]
    [Google Scholar]
  26. Veith A., Botelho H. M., Kindinger F., Gomes C. M., Kletzin A..( 2012;). The sulfur oxygenase reductase from the mesophilic bacterium Halothiobacillus neapolitanus is a highly active thermozyme. J Bacteriol194:677–685 [CrossRef][PubMed]
    [Google Scholar]
  27. Vesey C. J., Wilson J..( 1978;). Red cell cyanide. J Pharm Pharmacol30:20–26 [CrossRef][PubMed]
    [Google Scholar]
  28. Wood J. L..( 1975;). Biochemistry. Chemistry and Biochemistry of Thiocyanic Acid and its Derivatives156–221 Newman A. A.. London: Academic Press;
    [Google Scholar]
  29. Yamasaki M., Matsushita Y., Namura M., Nyunoya H., Katayama Y..( 2002;). Genetic and immunochemical characterization of thiocyanate-degrading bacteria in lake water. Appl Environ Microbiol68:942–946 [CrossRef][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.063339-0
Loading
/content/journal/micro/10.1099/mic.0.063339-0
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error