Molecular cloning of a copper-dependent laccase from the dye-decolorizing strain Stenotrophomonas maltophilia AAP56

Abstract

Aims: To clone the gene encoding the enzyme with laccase activity expressed by Stenotrophomonas maltophilia AAP56 and to construct an insertional mutation in that gene to determine its effect on dye decolourization and laccase activity in this strain.

Methods and results: Comparative genomics of Sten. maltophilia strains K279a and R551-3 revealed copA (coding for putative multicopper oxidase) as a candidate gene to encode an enzyme with laccase activity. Stenotrophomonas maltophilia AAP56 copA was amplified by degenerated PCR and cloned. A copA mutant strain, named Stemur, was constructed by homologous recombination. The comparison of wild-type and mutant strains revealed that CopA shows laccase activity, and it is involved in copper resistance and in vitro dye decolorization. On the contrary, the mutation in copA did not affect the in vivo dye removal by Sten. maltophilia.

Conclusions: Stenotrophomonas maltophilia AAP56 shows different mechanisms for dye decolorization. The gene encoding the laccase has been identified, and it has been shown that it is involved in the in vitro decolorization.

Significance and impact of the study: Stenotrophomonas maltophilia is a micro-organism of interest in different biotechnological processes including dye removal. This is the first report to address the molecular mechanism of this capacity, what will contribute to further improvements in the process.