Purification and characterization of two forms of endo-beta-1,4-mannanase from a thermotolerant fungus, Aspergillus fumigatus IMI 385708 (formerly Thermomyces lanuginosus IMI 158749)

Date

2004-11-01

Author

Puchart, V
Vrsanska, M
Svoboda, P
Pohl, J
Ögel, Zümrüt Begüm
Biely, P

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Two extracellular endo-β-1,4-mannanases, MAN I (major form) and MAN II (minor form), were purified to electrophoretic homogeneity from a locust bean gum-spent culture fluid of Aspergillus fumigatus IMI 385708 (formerly Thermomyces lanuginosus IMI 158749). Molecular weights of MAN I and MAN II estimated by SDS-PAGE were 60 and 63 kDa, respectively. IEF afforded several glycoprotein bands with pI values in the range of 4.9–5.2 for MAN I and 4.75–4.9 for MAN II, each exhibiting enzyme activity. MAN I as well as MAN II showed highest activity at pH 4.5 and 60 °C and were stable in the pH range 4.5–8.5 and up to 55 °C. In accordance with the ability of the enzymes to catalyze transglycosylation reactions, 1H NMR spectroscopy of reaction products generated from mannopentaitol confirmed the retaining character of both enzymes. Both MAN I and MAN II exhibited essentially identical kinetic parameters for polysaccharides and a similar hydrolysis pattern of various oligomeric and polymeric substrates. Both β-mannanases contained identical internal amino acid sequence corresponding to glycoside hydrolase family 5 and also a cellulose-binding module. These data suggested that both MAN I and MAN II are products of the same gene differing in posttranslational modification. Indeed, the corresponding gene was identified within the recently sequenced Aspergillus fumigatus genome (http://www.sanger.ac.uk/Projects/A_fumigatus/).

Subject Keywords

Biophysics, Biochemistry, Molecular Biology

URI

https://hdl.handle.net/11511/53404

Journal

BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS

DOI

https://doi.org/10.1016/j.bbagen.2004.06.022

Collections

Graduate School of Natural and Applied Sciences, Article

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Citation Formats

V. Puchart, M. Vrsanska, P. Svoboda, J. Pohl, Z. B. Ögel, and P. Biely, “Purification and characterization of two forms of endo-beta-1,4-mannanase from a thermotolerant fungus, Aspergillus fumigatus IMI 385708 (formerly Thermomyces lanuginosus IMI 158749),” BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS, pp. 239–250, 2004, Accessed: 00, 2020. [Online]. Available: https://hdl.handle.net/11511/53404.