tailieunhanh - Báo cáo khoa học: Role of the C-terminal extension in a bacterial tyrosinase Michael Fairhead and Linda Thony-Meyer

The well studied bacterial tyrosinases from theStreptomycessp. bacteria are distinguishable from their eukaryotic counterparts by the absence of a C-terminal extension. In the present study, we report that the tyrosinase from the bacterium Verrucomicrobium spinosumalso has such a C-terminal extension, thus making it distinct from the Streptomycesenzymes. | ỊFEBS Journal Role of the C-terminal extension in a bacterial tyrosinase Michael Fairhead and Linda Thony-Meyer EMPA Swiss FederalLaboratories for Materials Testing and Research Laboratory for Biomaterials St Gallen Switzerland Keywords C-terminaldomain melanin tyrosinase Verrucomicrobium spinosum zymogen Correspondence L. Thony-Meyer EMPA Swiss Federal Laboratories for Materials Testing and Research Laboratory for Biomaterials Lerchenfeldstrasse 5 St Gallen CH-9014 Switzerland Fax 41 44 071 274 7788 Tel 41 44 071 274 7792 E-mail Received 22 October 2009 revised 13 January 2010 accepted 22 February 2010 doi The well studied bacterial tyrosinases from the Streptomyces sp. bacteria are distinguishable from their eukaryotic counterparts by the absence of a C-terminal extension. In the present study we report that the tyrosinase from the bacterium Verrucomicrobium spinosum also has such a C-terminal extension thus making it distinct from the Streptomyces enzymes. The entire tyrosinase gene from V. spinosum codes for a 57 kDa protein fulllength unprocessed form which has a twin arginine translocase type signal peptide the two copper-binding motifs typical of the tyrosinase protein family and the aforementioned C-terminal extension. We expressed various mutants of the recombinant enzyme in Escherichia coli and found that removal of the C-terminal extension by genetic engineering or limited trypsin digest of the pro-form results in a more active enzyme . 30-100-fold increase in monophenolase and diphenolase activities . Further studies also revealed the importance of a phenylalanine residue in this C-terminal domain. These results demonstrate that the V. spinosum tyrosinase is a new example of this interesting family of enzymes. In addition we show that this enzyme can be readily overproduced and purified and that it will prove useful in furthering the understanding of these enzymes as well as their biotechnological