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Structural dynamics and catalytic properties of a multi-modular xanthanase
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)