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Applied and Environmental Microbiology, June 2004, p. 3609-3617, Vol. 70, No. 6
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.6.3609-3617.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Unusual Microbial Xylanases from Insect Guts

YaLi Brennan,¹ Walter N. Callen,¹ Leif Christoffersen,¹ Paul Dupree,² Florence Goubet,² Shaun Healey,¹ Myrian Hernández,³ Martin Keller,¹ Ke Li,¹ Nisha Palackal,¹ Ana Sittenfeld,⁴ Giselle Tamayo,^3,5 Steve Wells,¹ Geoffrey P. Hazlewood,¹ Eric J. Mathur,¹ Jay M. Short,¹ Dan E. Robertson,¹ and Brian A. Steer^1*

Diversa Corporation, San Diego, California 92121,¹ UEA-Bioprospección, Instituto Nacional de Biodiversidad, Santo Domingo, Heredia,³ Centro Investigación en Biología Celular y Molecular,⁴ Escuela de Química, Universidad de Costa Rica, San Pedro, Costa Rica,⁵ Department of Biochemistry, University of Cambridge, Cambridge CB2 1QW, United Kingdom²

Received 12 December 2003/ Accepted 3 March 2004

Recombinant DNA technologies enable the direct isolation and expression of novel genes from biotopes containing complex consortia of uncultured microorganisms. In this study, genomic libraries were constructed from microbial DNA isolated from insect intestinal tracts from the orders Isoptera (termites) and Lepidoptera (moths). Using a targeted functional assay, these environmental DNA libraries were screened for genes that encode proteins with xylanase activity. Several novel xylanase enzymes with unusual primary sequences and novel domains of unknown function were discovered. Phylogenetic analysis demonstrated remarkable distance between the sequences of these enzymes and other known xylanases. Biochemical analysis confirmed that these enzymes are true xylanases, which catalyze the hydrolysis of a variety of substituted ß-1,4-linked xylose oligomeric and polymeric substrates and produce unique hydrolysis products. From detailed polyacrylamide carbohydrate electrophoresis analysis of substrate cleavage patterns, the xylan polymer binding sites of these enzymes are proposed.

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* Corresponding author. Mailing address: Diversa Corporation, 4955 Directors Pl., San Diego, CA 92121. Phone: (800) 523-2990. Fax: (858) 526-5764. E-mail: bsteer{at}diversa.com.

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Applied and Environmental Microbiology, June 2004, p. 3609-3617, Vol. 70, No. 6
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.6.3609-3617.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

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