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Cloning, Isolation, and Properties of a New Recombinant Tannase from the Aspergillus niger Fungus

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Abstract

A new recombinant Aspergillus niger tannase (tannin acyl hydrolase) produced by the Penicillium verruculosum fungus has been studied. A strain with a high level of extracellular tannase (TAN2) secretion (80% of the total extracellular protein) was obtained by cloning the tan2 gene (PDB Acc. no: MT828303) into the recipient strain. The tannase enzyme preparation degraded tannins in black tea extracts. TAN2 was isolated in homogeneous form using chromatographic methods; the enzyme had a high activity with respect to gallotannin (53 U/mg) and less activity against propyl gallate (4.7 U/mg). Homogeneous TAN2 showed temperature and pH optima of 45°C and 3.5, respectively. At a temperature of 50°C, TAN2 retained above 80% of its activity for 3 h; at 60°C about 75% of its activity for 90 min was preserved; at 70°C, the enzyme was completely inactivated within 10 min. Tannase was characterized by a high tolerance to NaCl, the activity with respect to gallotannin exceeded 50% of the initial value in solutions with a salt concentration of up to 5 M. The tannase activity was stimulated by Ca2+, Mg2+, Zn2+, Mn2+, Cu2+, Cd2+, and Pb2+ by 3–64%, and inhibited by 4‒65% in the presence of Co2+, Fe3+ and Fe2+ ions.

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Funding

This work was supported by the Ministry of Science and Higher Education of the Russian Federation.

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Authors and Affiliations

  1. Fundamentals of Biotechnology Federal Research Center, Russian Academy of Sciences, 119071, Moscow, Russia

    D. O. Osipov, A. M. Rozhkova, I. A. Shashkov, A. D. Satrutdinov, E. G. Kondratyeva & A. P. Sinitsyn

  2. Skryabin Institute of Biochemistry and Physiology of Microorganisms, Russian Academy of Sciences, 142290, Pushchino, Russia

    V. Yu. Matys & V. A. Nemashkalov

  3. Moscow State University, 119991, Moscow, Russia

    A. P. Sinitsyn

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Abbreviations: BSA, bovine serum albumin; CBH1, cellobiohydrolase 1; CBH2, cellobiohydrolase 2; CL, culture liquid; EP, enzyme preparation; MCC, microcrystalline cellulose; OD520, optical density at a wavelength of 520 nm; TAN2, spray-dried tannase preparation.

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Osipov, D.O., Matys, V.Y., Nemashkalov, V.A. et al. Cloning, Isolation, and Properties of a New Recombinant Tannase from the Aspergillus niger Fungus. Appl Biochem Microbiol 58, 958–965 (2022). https://doi.org/10.1134/S000368382209006X

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