title: A family 11 xylanase from the pathogen Botrytis cinerea is inhibited by plant endoxylanase inhibitors XIP-I and TAXI-I.
abstract: The phytopathogen fungus Botrytis cinerea produces various glycosidases which are secreted during plant infection. In this study, the XynBc1 cDNA that encodes a xylanase from family 11 glycoside hydrolase from B. cinerea was identified by homology-based analysis, cloned by reverse transcription RT-PCR, fully sequenced, and heterologously expressed in Pichia pastoris X-33. The purified recombinant protein obtained by chelating-affinity chromatography demonstrated high catalytic activity (180+/-23 U/mg) and efficiently degraded low viscosity xylan [K(m) = 10+/-3 g L(-1), V(max) = 0.50+/-0.04 micromol xylose min(-1), and k(cat) = 136+/-11.5 s(-1) at pH 4.5 and 25 degrees C]. XynBc1 was further tested for its ability to interact with wheat XIP and TAXI type xylanase inhibitors which have been implicated in plant defence. The xylanase activity of XynBc1 produced in P. pastoris was strongly inhibited by both XIP-I and TAXI-I in a competitive manner, with a K(i) of 2.1+/-0.1 and 6.0+/-0.2 nM, respectively, whereas no inhibition was detected with TAXI-II. We also showed that XynBc1 mRNAs accumulated during early stages of plant tissue infection.
authors.affiliation: Institut Méditerranéen de Recherche en Nutrition, Laboratoire de Biochimie et Biologie de la Nutrition, UMR Université Paul Cézanne Aix Marseille III, INRA 1111, service 342, Faculté des Sciences et Techniques Saint-Jérôme, Marseille Cedex, France.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Brutus
authors.forename: Ida Barbara
authors.initials: IB
authors.lastname: Reca
authors.forename: Sameh
authors.initials: S
authors.lastname: Herga
authors.forename: Benedetta
authors.initials: B
authors.lastname: Mattei
authors.forename: Antoine
authors.initials: A
authors.lastname: Puigserver
authors.forename: Jean-Claude
authors.initials: JC
authors.lastname: Chaix
authors.forename: Nathalie
authors.initials: N
authors.lastname: Juge
authors.forename: Daniela
authors.initials: D
authors.lastname: Bellincampi
authors.forename: Thierry
authors.initials: T
authors.lastname: Giardina
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D020171:Botrytis / Q000201:enzymology*; D002352:Carrier Proteins / Q000494:pharmacology*; D003001:Cloning, Molecular; D043364:Endo-1,4-beta Xylanases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins; D008969:Molecular Sequence Data; D010843:Pichia / Q000235:genetics; D010940:Plant Proteins / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000378:metabolism; D016415:Sequence Alignment
chemical_list: D002352:Carrier Proteins; D004791:Enzyme Inhibitors; D005656:Fungal Proteins; D047908:Intracellular Signaling Peptides and Proteins; D010940:Plant Proteins; D011994:Recombinant Proteins; C422246:TAXI I protein, Triticum aestivum; C485063:XIP-I protein, Triticum aestivum; D043364:Endo-1,4-beta Xylanases