Molecular cloning and enzymatic characterization of a Trichoderma reesei 1,2-α-D-mannosidase

Marleen Maras, Nico Callewaert, Kathleen Piens, Marc Claeyssens, Wim Martinet, Sylviane Dewaele, Hans Contreras, Isabelle Dewerte, Merja Penttilä, Roland Contreras

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Abstract

A cDNA encoding 1,2-α-D-mannosidase mds1 from Trichoderma reesei was cloned. The largest open reading frame occupied 1571 bp. The predicted sequence contains 523 amino acid residues for a calculated molecular mass of 56 266 Da and shows high similarity to the amino acid sequences of 1,2-α-D-mannosidases from Aspergillus saitoi and Penicillium citrinum (51.6 and 51.0% identity, respectively). T. reesei mannosidase was produced as a recombinant enzyme in the yeast Pichia pastoris. Replacement of the N-terminal part with the prepro-signal peptide of the Saccharomyces cerevisiae α-mating factor resulted in high amounts of secreted enzyme. A three-step purification protocol was designed and the enzymatic properties were analysed. The enzyme was characterized as a class-I mannosidase. Copyright (C) 2000 Elsevier Science B.V.

Original languageEnglish
Pages (from-to)255-263
Number of pages9
JournalJournal of Biotechnology
Volume77
Issue number2-3
DOIs
Publication statusPublished - 17 Feb 2000
MoE publication typeA1 Journal article-refereed

Keywords

  • Fungus
  • Glycosidase
  • Mannosidase
  • Oligosaccharide
  • Trichoderma reesei
  • Yeast

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    Maras, M., Callewaert, N., Piens, K., Claeyssens, M., Martinet, W., Dewaele, S., Contreras, H., Dewerte, I., Penttilä, M., & Contreras, R. (2000). Molecular cloning and enzymatic characterization of a Trichoderma reesei 1,2-α-D-mannosidase. Journal of Biotechnology, 77(2-3), 255-263. https://doi.org/10.1016/S0168-1656(99)00222-9