Current Genetics

, Volume 30, Issue 6, pp 531–540

Molecular characterization of xyn3, a member of the endoxylanase multigene family of the rumen anaerobic fungus Neocallimastix frontalis

  • R. Durand
  • Christine Rascle
  • Michel Fèvre
ORIGINAL PAPER

DOI: 10.1007/s002940050166

Cite this article as:
Durand, R., Rascle, C. & Fèvre, M. Curr Genet (1996) 30: 531. doi:10.1007/s002940050166

Abstract

Different cDNAs designated xyn3 and xyn4 were isolated from an expression library of the anaerobic rumen fungus Neocallimastix frontalis. Xyn3 was further characterized and was shown to contain a single open reading frame of 1821 bp coding for a protein, XYN3, of 607 amino acids (Mr 66 000). The predicted primary structure of XYN3 consisted of two large reiterated regions of 223 amino acids with a high degree of identity (88.3%). Each domain of XYN3, XYN3A and XYN3B, showed significant homology with fungal and bacterial xylanases belonging to the endoxylanase family 11. XYN3 and XYN3A were cloned in a bacterial expression plasmid harbouring a 6 His-C terminal tag and the recombinant proteins XYN3 and XYN3A were purified from Escherichia coli. The recombinant proteins had Mr of 66 800 and 34 000 respectively and hydrolysed xylan to xylo-oligosaccharides. Analysis of truncated forms of XYN3 confirmed that the full-length protein contained two catalytic domains displaying similar substrate specificity. Western-blot analysis using antiserum raised against XYN3A showed that the N. frontalis xylanase was not submitted to post-translational maturation. XYN3A antiserum recognized similar polypeptides in the culture medium of two other rumen fungi, Piromyces rhizinflata and Caecomyces communis.

Key words Anaerobic fungus Neocallimastix frontalis Endoxylanase cDNA Bacterial recombinant xylanases 

Copyright information

© Springer-Verlag Berlin Heidelberg 1996

Authors and Affiliations

  • R. Durand
    • 1
  • Christine Rascle
    • 1
  • Michel Fèvre
    • 1
  1. 1.Laboratoire de Biologie Cellulaire Fongique, Centre de Génétique Moléculaire et Cellulaire, CNRS UMR 5534, Bât 405, Université Claude Bernard Lyon I, F-69622 Villeurbanne Cedex, FranceFR

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