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Structural dynamics and catalytic properties of a multi-modular xanthanase

DOI: 10.1021/acscatal.8b00666 DOI Help

Authors: Olga V. Moroz (University of York) , Pernille Foged Jensen (University of Copenhagen) , Sean P Mcdonald (University of British Columbia) , Nicholas Mcgregor (University of British Columbia) , Elena Blagova (The University of York) , Gerard Comamala (University of Copenhagen) , Dorotea R. Segura (Novozymes A/S) , Lars Anderson (Novozymes A/S) , Santhosh M Vasu (Novozymes A/S) , Vasudeva P Rao (Novozymes A/S) , Lars Giger (Novozymes A/S) , Trine Holst Sørensen (Roskilde University) , Rune Nygaard Monrad (Novozymes A/S) , Allan Svendsen (Novozymes A/S) , Jens Erik Nielsen (Novozymes A/S) , Bernard Henrissat (Aix-Marseille University; King Abdulaziz University) , Gideon Davies (The University of York) , Harry Brumer (University of British Columbia) , Kasper D. Rand (University of Copenhagen) , Keith S. Wilson (The University of York)
Co-authored by industrial partner: Yes

Type: Journal Paper
Journal: Acs Catalysis

State: Published (Approved)
Published: May 2018
Diamond Proposal Number(s): 7864

Abstract: The precise catalytic strategies used for the breakdown of the complex bacterial polysaccharide xanthan, an increasingly frequent component of processed human foodstuffs, have remained a mystery. Here we present the characterization of an endo-xanthanase from Paenibacillus sp. 62047. We show that it is a CAZy family 9 glycoside hydrolase (GH9) responsible for the hydrolysis of the xanthan backbone, capable of generating tetrameric xanthan oligosaccharides from polysaccharide lyase family 8 (PL8) xanthan lyase-treated xanthan. 3-D structure determination reveals a complex multi-modular enzyme in which a catalytic (α/α)6 barrel is flanked by an N-terminal "immunoglobulin-like" (Ig-like) domain (frequently found in GH9 enzymes) and by four additional C-terminal all β-sheet domains which have very few homologs in sequence databases and, at least, one of which functions as a new xanthan-binding domain, now termed CBM84. The solution phase conformation and dynamics of the enzyme in the native calcium-bound state and in the absence of calcium were probed experimentally by hydrogen/deuterium exchange mass spectrometry. Measured conformational dynamics were used to guide the protein engineering of enzyme variants with increased stability in the absence of calcium; a property of interest for the potential use of the enzyme in cleaning detergents. The ability of hydrogen/deuterium exchange mass spectrometry to pinpoint dynamic regions of a protein under stress (e.g. removal of calcium ions) makes this technology a strong tool for improving protein catalyst properties by informed engineering.

Journal Keywords: Enzyme; carbohydrate; Xanthan; Hydrogen/deuterium exchange mass spectrometry; enzyme stability; enzyme dynamics

Subject Areas: Biology and Bio-materials, Chemistry


Instruments: I03-Macromolecular Crystallography , I04-1-Macromolecular Crystallography (fixed wavelength)