A hyperthermophilic laccase from Thermus thermophilus HB27
- 6 July 2005
- journal article
- Published by Springer Science and Business Media LLC in Extremophiles
- Vol. 9 (6), 415-425
- https://doi.org/10.1007/s00792-005-0458-z
Abstract
A copper-inducible laccase activity was detected in Thermus thermophilus HB27. The enzyme was partially purified and separated by SDS-PAGE. After staining, a gel slice containing a ~53-kDa protein was excised and treated with trypsin, and the in-gel digests were analyzed by mass spectrometry. By mass fingerprinting, the peptides were found to share identity with the TTC1370 protein of the thermophile, which was tentatively annotated as a laccase in the whole genome analysis, albeit experimental evidence was lacking. The assigned mass nearest to the N-terminal sequence was that from Gln23 to Lys31. By signal peptide prediction, TTC1370 protein was assumed to be a secretory protein starting from Gln23. The DNA encoding the mature protein was then cloned and expressed in Escherichia coli. The recombinant enzyme, expressed as an apoprotein, was dialyzed against copper-containing buffer to yield a holoprotein. The holoprotein was purified to homogeneity, which displayed a blue color typical of laccases and oxidized canonical laccase substrates such as guaiacol and 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonate). The enzyme was most notable for its striking thermophilicity; the optimal reaction temperature was ~92°C and the half-life of thermal inactivation at 80°C was >14 h, ranking it as the most thermophilic laccase reported thus far.Keywords
This publication has 34 references indexed in Scilit:
- Improved Prediction of Signal Peptides: SignalP 3.0Journal of Molecular Biology, 2004
- Role of oxidative enzymatic treatments on enzymatic hydrolysis of softwoodBiotechnology & Bioengineering, 2004
- The genome sequence of the extreme thermophile Thermus thermophilusNature Biotechnology, 2004
- White-rot fungi and their enzymes for the treatment of industrial dye effluentsBiotechnology Advances, 2003
- Redox-mediated decolorization of synthetic dyes by fungal laccasesApplied Microbiology and Biotechnology, 2002
- Laccase: new functions for an old enzymePhytochemistry, 2002
- CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choiceNucleic Acids Research, 1994
- Basic local alignment search toolJournal of Molecular Biology, 1990
- Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genesJournal of Molecular Biology, 1986
- Calculation of the isoelectric points of polypeptides from the amino acid compositionTrAC Trends in Analytical Chemistry, 1986