Potential biological role of laccase from the sponge Suberites domuncula as an antibacterial defense component

• Published in: Biochimica et biophysica acta Vol. 1850; no. 1; pp. 118 - 128
• Main Authors: Li, Qiang, Wang, Xiaohong, Korzhev, Michael, Schröder, Heinz C., Link, Thorben, Tahir, Muhammad Nawaz, Diehl-Seifert, Bärbel, Müller, Werner E.G.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01-01-2015
• Subjects: Amino Acid Sequence; Animals; Anti-Bacterial Agents - metabolism; Anti-bacterial defense; Biocatalysis; Copper; Dose-Response Relationship, Drug; Escherichia coli - drug effects; Escherichia coli - growth & development; Ferric Compounds - chemistry; Ferromagnetic particles; Gene Expression Regulation, Enzymologic - drug effects; Hydrazones - metabolism; Kinetics; Laccase; Laccase - classification; Laccase - genetics; Laccase - metabolism; Lignin; Lignin - metabolism; Lipopolysaccharides - pharmacology; Molecular Sequence Data; Nanoparticles - chemistry; Nanoparticles - toxicity; Oxidation-Reduction; Phylogeny; Recombinant Proteins - metabolism; Reverse Transcriptase Polymerase Chain Reaction; Sequence Homology, Amino Acid; Sponges; Suberites - enzymology; Suberites - genetics; Substrate Specificity; Up-Regulation - drug effects; Ferromagnetic particles; Lignin; Sponges; Anti-bacterial defense; Copper; Laccase
• ISSN: 0304-4165; 0006-3002; 1872-8006
• DOI: 10.1016/j.bbagen.2014.10.007

Laccases are copper-containing enzymes that catalyze the oxidation of a wide variety of phenolic substrates.
We describe the first poriferan laccase from the marine demosponge Suberites domuncula.
This enzyme comprises three characteristic multicopper oxidase homologous domains. Immunohistological studies revealed that the highest expression of the laccase is in the surface zone of the animals. The expression level of the laccase gene is strongly upregulated after exposure of the animals to the bacterial endotoxin lipopolysaccharide. To allow the binding of the recombinant enzyme to ferromagnetic nanoparticles, a recombinant laccase was prepared which contained in addition to the His-tag, a Glu-tag at the N-terminus of the enzyme. The recombinant laccase was enzymatically active. The apparent Michaelis constant of the enzyme is 114μM, using syringaldazine as substrate. Exposure of E. coli to the nanoparticles, coated with Glu-tagged laccase, and to the mediator 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) in the presence of lignin, as the oxidizable substrate, resulted in an almost complete inhibition of colony formation. Quantitative studies of the effect of the laccase-coated iron oxide nanoparticles were performed using E. coli grown in suspension in reaction tubes within a magnetic nanoparticle separator.
This newly designed magnetic nanoparticle separator allowed a removal of the nanoparticles after terminating the reaction. Using this system, a strong dose-dependent inhibition of the growth of E. coli by the laccase iron oxide nanoparticles was determined.
From our data we conclude that the sponge laccase is involved in the anti-bacterial defense of the sponge organism.
A siliceous sponge (Suberites domuncula) comprises a laccase [Lac] that functions as a radical generating enzyme by which bacteria are killed. In order to enhance the function the enzyme employs a mediator, e.g. ABTS. [Display omitted]
•Sponges, the oldest Metazoa, are exposed to a heavy load of bacteria and viruses.•They comprise a laccase, catalyzing the oxidation of substrates via free radical mechanisms.•We propose that these animals can kill environmental bacteria by radical formation — a new defense mechanism for them.