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Using neutron crystallography to elucidate the basis of selective inhibition of carbonic anhydrase by saccharin and a derivative

Koruza, Katarina LU ; Mahon, Brian P.; Blakeley, Matthew P.; Ostermann, Andreas; Schrader, Tobias E.; McKenna, Robert; Knecht, Wolfgang LU and Fisher, S. Zoë LU (2019) In Journal of Structural Biology
Abstract

Up-regulation of carbonic anhydrase IX (CA IX) expression is an indicator of metastasis and associated with poor cancer patient prognosis. CA IX has emerged as a cancer drug target but development of isoform-specific inhibitors is challenging due to other highly conserved CA isoforms. In this study, a CA IXmimic construct was used (CA II with seven point mutations introduced, to mimic CA IX active site) while maintaining CA II solubility that make it amenable to crystallography. The structures of CA IXmimic unbound and in complex with saccharin (SAC) and a saccharin-glucose conjugate (SGC) were determined using joint X-ray and neutron protein crystallography. Previously, SAC and SGC have been shown to display CA... (More)

Up-regulation of carbonic anhydrase IX (CA IX) expression is an indicator of metastasis and associated with poor cancer patient prognosis. CA IX has emerged as a cancer drug target but development of isoform-specific inhibitors is challenging due to other highly conserved CA isoforms. In this study, a CA IXmimic construct was used (CA II with seven point mutations introduced, to mimic CA IX active site) while maintaining CA II solubility that make it amenable to crystallography. The structures of CA IXmimic unbound and in complex with saccharin (SAC) and a saccharin-glucose conjugate (SGC) were determined using joint X-ray and neutron protein crystallography. Previously, SAC and SGC have been shown to display CA isoform inhibitor selectivity in assays and X-ray crystal structures failed to reveal the basis of this selectivity. Joint X-ray and neutron crystallographic studies have shown active site residues, solvent, and H-bonding re-organization upon SAC and SGC binding. These observations highlighted the importance of residues 67 (Asn in CA II, Gln in CA IX) and 130 (Asp in CA II, Arg in CA IX) in selective CA inhibitor targeting.

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author
organization
publishing date
type
Contribution to journal
publication status
epub
subject
keywords
Crystallography, H/D exchange, Hydrogen atoms, Joint X-ray neutron refinement, Ligand protein interactions
in
Journal of Structural Biology
publisher
Elsevier
external identifiers
  • scopus:85060113121
ISSN
1047-8477
DOI
10.1016/j.jsb.2018.12.009
language
English
LU publication?
yes
id
bd5ba350-73e6-48d7-8339-5c7831a2c4d2
date added to LUP
2019-01-29 14:25:41
date last changed
2019-02-20 11:45:12
@article{bd5ba350-73e6-48d7-8339-5c7831a2c4d2,
  abstract     = {<p>Up-regulation of carbonic anhydrase IX (CA IX) expression is an indicator of metastasis and associated with poor cancer patient prognosis. CA IX has emerged as a cancer drug target but development of isoform-specific inhibitors is challenging due to other highly conserved CA isoforms. In this study, a CA IX<sub>mimic</sub> construct was used (CA II with seven point mutations introduced, to mimic CA IX active site) while maintaining CA II solubility that make it amenable to crystallography. The structures of CA IX<sub>mimic</sub> unbound and in complex with saccharin (SAC) and a saccharin-glucose conjugate (SGC) were determined using joint X-ray and neutron protein crystallography. Previously, SAC and SGC have been shown to display CA isoform inhibitor selectivity in assays and X-ray crystal structures failed to reveal the basis of this selectivity. Joint X-ray and neutron crystallographic studies have shown active site residues, solvent, and H-bonding re-organization upon SAC and SGC binding. These observations highlighted the importance of residues 67 (Asn in CA II, Gln in CA IX) and 130 (Asp in CA II, Arg in CA IX) in selective CA inhibitor targeting.</p>},
  author       = {Koruza, Katarina and Mahon, Brian P. and Blakeley, Matthew P. and Ostermann, Andreas and Schrader, Tobias E. and McKenna, Robert and Knecht, Wolfgang and Fisher, S. Zoë},
  issn         = {1047-8477},
  keyword      = {Crystallography,H/D exchange,Hydrogen atoms,Joint X-ray neutron refinement,Ligand protein interactions},
  language     = {eng},
  month        = {01},
  publisher    = {Elsevier},
  series       = {Journal of Structural Biology},
  title        = {Using neutron crystallography to elucidate the basis of selective inhibition of carbonic anhydrase by saccharin and a derivative},
  url          = {http://dx.doi.org/10.1016/j.jsb.2018.12.009},
  year         = {2019},
}