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Characterization of a multi-tolerant tannin acyl hydrolase II from Aspergillus carbonarius produced under solid-state fermentation

Valera, Larissa Serrani, Jorge, João Atílio, Guimarães, Luis Henrique Souza
Electronic Journal of Biotechnology 2015 v.18 pp. 464-470
Aspergillus carbonarius, activation energy, agarose, biotechnology, carbohydrate content, carbon, detergents, enzyme activity, fungi, gel chromatography, glycoproteins, green tea, half life, leaves, pH, polyacrylamide gel electrophoresis, silver nitrate, solid state fermentation, solvents, tannase, tannins, tap water, temperature
Tannases are enzymes with biotechnological potential produced mainly by microorganisms as filamentous fungi. In this context, the production and characterization of a multi-tolerant tannase from Aspergillus carbonarius is described.The filamentous fungus A. carbonarius produced high levels of tannase when cultivated under solid-state fermentation using green tea leaves as substrate/carbon source and tap water at a 1:1 ratio as the moisture agent for 72h at 30°C. Two tannase activity peaks were obtained during the purification step using DEAE-Cellulose. The second peak (peak II) was purified 11-fold with 14% recovery from a Sepharose CL-6B chromatographic column. The tannase from peak II (tannase II) was characterized as a heterodimeric glycoprotein of 134.89kDa, estimated through gel filtration, with subunits of 65kDa and 100kDa, estimated through SDS-PAGE, and 48% carbohydrate content. The optimal temperature and pH for tannase II activity was 60°C and 5.0, respectively. The enzyme was fully stable at temperatures ranging from 20–60°C for 120min, and the half-life (T1/2) at 75°C was 62min. The activation energy was 28.93kJ/mol. After incubation at pH5.0 for 60min, 75% of the enzyme activity was maintained. However, enzyme activity was increased in the presence of AgNO3 and it was tolerant to solvents and detergents. Tannase II exhibited a better affinity for methyl gallate (Km=1.42mM) rather than for tannic acid (Km=2.2mM).A. carbonarius tannase presented interesting properties as, for example, multi-tolerance, which highlight its potential for future application.