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Crystallization, neutron data collection, initial structure refinement and analysis of a xyloglucan heptamer bound to an engineered carbohydrate-binding module from xylanase.

Ohlin, Mats LU ; von Schantz, Laura LU ; Schrader, Tobias E; Ostermann, Andreas; Logan, Derek LU and Fisher, S Zoë (2015) In Acta crystallographica. Section F, Structural biology communications 71(Pt 8). p.1072-1077
Abstract
Carbohydrate-binding modules (CBMs) are discrete parts of carbohydrate-hydrolyzing enzymes that bind specific types of carbohydrates. Ultra high-resolution X-ray crystallographic studies of CBMs have helped to decipher the basis for specificity in carbohydrate-protein interactions. However, additional studies are needed to better understand which structural determinants confer which carbohydrate-binding properties. To address these issues, neutron crystallographic studies were initiated on one experimentally engineered CBM derived from a xylanase, X-2 L110F, a protein that is able to bind several different plant carbohydrates such as xylan, β-glucan and xyloglucan. This protein evolved from a CBM present in xylanase Xyn10A of Rhodothermus... (More)
Carbohydrate-binding modules (CBMs) are discrete parts of carbohydrate-hydrolyzing enzymes that bind specific types of carbohydrates. Ultra high-resolution X-ray crystallographic studies of CBMs have helped to decipher the basis for specificity in carbohydrate-protein interactions. However, additional studies are needed to better understand which structural determinants confer which carbohydrate-binding properties. To address these issues, neutron crystallographic studies were initiated on one experimentally engineered CBM derived from a xylanase, X-2 L110F, a protein that is able to bind several different plant carbohydrates such as xylan, β-glucan and xyloglucan. This protein evolved from a CBM present in xylanase Xyn10A of Rhodothermus marinus. The protein was complexed with a branched xyloglucan heptasaccharide. Large single crystals of hydrogenous protein (∼1.6 mm(3)) were grown at room temperature and subjected to H/D exchange. Both neutron and X-ray diffraction data sets were collected to 1.6 Å resolution. Joint neutron and X-ray refinement using phenix.refine showed significant density for residues involved in carbohydrate binding and revealed the details of a hydrogen-bonded water network around the binding site. This is the first report of a neutron structure of a CBM and will add to the understanding of protein-carbohydrate binding interactions. (Less)
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Contribution to journal
publication status
published
subject
in
Acta crystallographica. Section F, Structural biology communications
volume
71
issue
Pt 8
pages
1072 - 1077
publisher
Wiley-Blackwell
external identifiers
  • pmid:26249702
  • wos:000359352700026
  • scopus:84938863810
ISSN
2053-230X
DOI
10.1107/S2053230X15011383
language
English
LU publication?
yes
id
a9f4959f-911b-4810-b748-1feccf7a9922 (old id 7844491)
date added to LUP
2015-09-10 10:50:18
date last changed
2017-07-30 04:16:42
@article{a9f4959f-911b-4810-b748-1feccf7a9922,
  abstract     = {Carbohydrate-binding modules (CBMs) are discrete parts of carbohydrate-hydrolyzing enzymes that bind specific types of carbohydrates. Ultra high-resolution X-ray crystallographic studies of CBMs have helped to decipher the basis for specificity in carbohydrate-protein interactions. However, additional studies are needed to better understand which structural determinants confer which carbohydrate-binding properties. To address these issues, neutron crystallographic studies were initiated on one experimentally engineered CBM derived from a xylanase, X-2 L110F, a protein that is able to bind several different plant carbohydrates such as xylan, β-glucan and xyloglucan. This protein evolved from a CBM present in xylanase Xyn10A of Rhodothermus marinus. The protein was complexed with a branched xyloglucan heptasaccharide. Large single crystals of hydrogenous protein (∼1.6 mm(3)) were grown at room temperature and subjected to H/D exchange. Both neutron and X-ray diffraction data sets were collected to 1.6 Å resolution. Joint neutron and X-ray refinement using phenix.refine showed significant density for residues involved in carbohydrate binding and revealed the details of a hydrogen-bonded water network around the binding site. This is the first report of a neutron structure of a CBM and will add to the understanding of protein-carbohydrate binding interactions.},
  author       = {Ohlin, Mats and von Schantz, Laura and Schrader, Tobias E and Ostermann, Andreas and Logan, Derek and Fisher, S Zoë},
  issn         = {2053-230X},
  language     = {eng},
  number       = {Pt 8},
  pages        = {1072--1077},
  publisher    = {Wiley-Blackwell},
  series       = {Acta crystallographica. Section F, Structural biology communications},
  title        = {Crystallization, neutron data collection, initial structure refinement and analysis of a xyloglucan heptamer bound to an engineered carbohydrate-binding module from xylanase.},
  url          = {http://dx.doi.org/10.1107/S2053230X15011383},
  volume       = {71},
  year         = {2015},
}