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Properties of a purified thermostable glucoamylase from Aspergillus niveus

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Journal of Industrial Microbiology & Biotechnology

Abstract

A glucoamylase from Aspergillus niveus was produced by submerged fermentation in Khanna medium, initial pH 6.5 for 72 h, at 40°C. The enzyme was purified by DEAE-Fractogel and Concanavalin A-Sepharose chromatography. The enzyme showed 11% carbohydrate content, an isoelectric point of 3.8 and a molecular mass of 77 and 76 kDa estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis or Bio-Sil-Sec-400 gel filtration, respectively. The pH optimum was 5.0–5.5, and the enzyme remained stable for at least 2 h in the pH range of 4.0–9.5. The temperature optimum was 65°C and retained 100% activity after 240 min at 60°C. The glucoamylase remained completely active in the presence of 10% methanol and acetone. After 120 min hydrolysis of starch, glucose was the unique product formed, confirming that the enzyme was a glucoamylase (1,4-alpha-d-glucan glucohydrolase). The K m was calculated as 0.32 mg ml−1. Circular dichroism spectroscopy estimated a secondary structure content of 33% α-helix, 17% β-sheet and 50% random structure, which is similar to that observed in the crystal structures of glucoamylases from other Aspergillus species. The tryptic peptide sequence analysis showed similarity with glucoamylases from A. niger, A. kawachi, A. ficcum, A. terreus, A. awamori and A. shirousami. We conclude that the reported properties, such as solvent, pH and temperature stabilities, make A. niveus glucoamylase a potentially attractive enzyme for biotechnological applications.

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Acknowledgments

This work was supported by grants from Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) and Conselho de Desenvolvimento Científico e Tecnológico (CNPQ). J. A. Jorge, H. F. Terenzi and M. L. T. M. Polizeli are Research Fellows of CNPq. T. M. Silva is recipient FAPESP Fellowship and this work was part of a Doctor Thesis submitted by T. M. Silva to FFCLRP-USP. We thank Ricardo F. Alarcon and Mauricio de Oliveira for technical assistance.

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Authors and Affiliations

  1. Departamento de Biologia, Faculdade de Filosofia Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Av. Bandeirantes 3900, Monte Alegre, 14040-901, Ribeirão Preto, São Paulo, Brazil

    Tony Marcio da Silva, Michele Michelin, João Atilio Jorge, Héctor Francisco Terenzi & Maria Lourdes T. M. de Polizeli

  2. Departamento de Bioquímica e Imunologia, FMRP, Universidade de São Paulo, Ribeirão Preto, São Paulo, Brazil

    Alexandre Maller & André Ricardo de Lima Damásio

  3. Departamento de Química, FFCLRP, Universidade de São Paulo, Ribeirão Preto, São Paulo, Brazil

    Richard John Ward

  4. Departamento de Biofísica, Universidade Federal de São Paulo, São Paulo, SP, Brazil

    Izaura Yoshico Hirata

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  1. Tony Marcio da Silva
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  2. Alexandre Maller
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  9. Maria Lourdes T. M. de Polizeli
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Correspondence to Maria Lourdes T. M. de Polizeli.

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da Silva, T.M., Maller, A., de Lima Damásio, A.R. et al. Properties of a purified thermostable glucoamylase from Aspergillus niveus . J Ind Microbiol Biotechnol 36, 1439–1446 (2009). https://doi.org/10.1007/s10295-009-0630-z

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  • DOI: https://doi.org/10.1007/s10295-009-0630-z

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