Unique features of the bifunctional GH30 from Thermothelomyces thermophila revealed by structural and mutational studies. (1st December 2021)

• Record Type: Journal Article
• Authors: Nikolaivits, Efstratios; Pentari, Christina; Kosinas, Christos; Feiler, Christian G.; Spiliopoulou, Maria; Weiss, Manfred S.; Dimarogona, Maria; Topakas, Evangelos
• Is Part Of: Carbohydrate polymers. Volume 273(2021)
• Publication Date: 2021-12-01
• Subjects: GH30 xylanase -- Xylobiohydrolase -- Acidic xylooligosaccharides -- Glucuronic acid recognition -- X-ray structure -- Protein engineering; Polysaccharides -- Periodicals; Polysaccharides -- Periodicals; Polysaccharides -- Périodiques; Electronic journals; 547.78
• DOI: 10.1016/j.carbpol.2021.118553
• Languages: English
• ISSNs: 0144-8617

Abstract

Abstract: Fungal xylanases belonging to family GH30_7, initially categorized as endo -glucuronoxylanases, are now known to differ both in terms of substrate specificity, as well as mode of action. Recently, Tt Xyn30A, a GH30_7 xylanase from Thermothelomyces thermophila, was shown to possess dual activity, acting on the xylan backbone in both an endo - and an exo - manner. Here, in an effort to identify the structural characteristics that append these functional properties to the enzyme, we present the biochemical characterization of various Tt Xyn30A mutants as well as its crystal structure, alone, and in complex with the reaction product. An auxiliary catalytic amino acid has been identified, while it is also shown that glucuronic acid recognition is not mediated by a conserved arginine residue, as shown by previously determined GH30 structures. Highlights: Determination of the second crystal structure of a GH30_7 xylanase Structural and mutational studies shed light on dual (endo/exo) enzyme function. An auxiliary catalytic glutamate was identified in the absence of the conserved one. Glucuronic acid recognition is not mediated by a conserved Arg as in other GH30s.