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Structural comparison of protiated, H/D-exchanged and deuterated human carbonic anhydrase IX

Koruza, K LU ; Lafumat, B LU ; Nyblom, M LU ; Mahon, B P ; Knecht, W LU ; McKenna, R and Fisher, S Z LU (2019) In Acta Crystallographica Section D: Structural Biology 75(Pt 10). p.895-903
Abstract

Human carbonic anhydrase IX (CA IX) expression is upregulated in hypoxic solid tumours, promoting cell survival and metastasis. This observation has made CA IX a target for the development of CA isoform-selective inhibitors. To enable structural studies of CA IX-inhibitor complexes using X-ray and neutron crystallography, a CA IX surface variant (CA IXSV; the catalytic domain with six surface amino-acid substitutions) has been developed that can be routinely crystallized. Here, the preparation of protiated (H/H), H/D-exchanged (H/D) and deuterated (D/D) CA IXSV for crystallographic studies and their structural comparison are described. Four CA IXSV X-ray crystal structures are compared: two H/H crystal forms, an H/D crystal form and a... (More)

Human carbonic anhydrase IX (CA IX) expression is upregulated in hypoxic solid tumours, promoting cell survival and metastasis. This observation has made CA IX a target for the development of CA isoform-selective inhibitors. To enable structural studies of CA IX-inhibitor complexes using X-ray and neutron crystallography, a CA IX surface variant (CA IXSV; the catalytic domain with six surface amino-acid substitutions) has been developed that can be routinely crystallized. Here, the preparation of protiated (H/H), H/D-exchanged (H/D) and deuterated (D/D) CA IXSV for crystallographic studies and their structural comparison are described. Four CA IXSV X-ray crystal structures are compared: two H/H crystal forms, an H/D crystal form and a D/D crystal form. The overall active-site organization in each version is essentially the same, with only minor positional changes in active-site solvent, which may be owing to deuteration and/or resolution differences. Analysis of the crystal contacts and packing reveals different arrangements of CA IXSV compared with previous reports. To our knowledge, this is the first report comparing three different deuterium-labelled crystal structures of the same protein, marking an important step in validating the active-site structure of CA IXSV for neutron protein crystallography.

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author
; ; ; ; ; and
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
Antigens, Neoplasm/chemistry, Carbonic Anhydrase IX/chemistry, Catalytic Domain, Cloning, Molecular, Crystallography, X-Ray/methods, Deuterium, Escherichia coli/genetics, Humans, Models, Molecular
in
Acta Crystallographica Section D: Structural Biology
volume
75
issue
Pt 10
pages
9 pages
publisher
John Wiley & Sons Inc.
external identifiers
  • scopus:85072919114
  • pmid:31588921
ISSN
2059-7983
DOI
10.1107/S2059798319010027
language
English
LU publication?
yes
additional info
open access.
id
3b68a104-156d-4e1b-9641-8f5c2263760f
date added to LUP
2020-07-09 16:33:31
date last changed
2024-05-30 19:14:17
@article{3b68a104-156d-4e1b-9641-8f5c2263760f,
  abstract     = {{<p>Human carbonic anhydrase IX (CA IX) expression is upregulated in hypoxic solid tumours, promoting cell survival and metastasis. This observation has made CA IX a target for the development of CA isoform-selective inhibitors. To enable structural studies of CA IX-inhibitor complexes using X-ray and neutron crystallography, a CA IX surface variant (CA IXSV; the catalytic domain with six surface amino-acid substitutions) has been developed that can be routinely crystallized. Here, the preparation of protiated (H/H), H/D-exchanged (H/D) and deuterated (D/D) CA IXSV for crystallographic studies and their structural comparison are described. Four CA IXSV X-ray crystal structures are compared: two H/H crystal forms, an H/D crystal form and a D/D crystal form. The overall active-site organization in each version is essentially the same, with only minor positional changes in active-site solvent, which may be owing to deuteration and/or resolution differences. Analysis of the crystal contacts and packing reveals different arrangements of CA IXSV compared with previous reports. To our knowledge, this is the first report comparing three different deuterium-labelled crystal structures of the same protein, marking an important step in validating the active-site structure of CA IXSV for neutron protein crystallography.</p>}},
  author       = {{Koruza, K and Lafumat, B and Nyblom, M and Mahon, B P and Knecht, W and McKenna, R and Fisher, S Z}},
  issn         = {{2059-7983}},
  keywords     = {{Antigens, Neoplasm/chemistry; Carbonic Anhydrase IX/chemistry; Catalytic Domain; Cloning, Molecular; Crystallography, X-Ray/methods; Deuterium; Escherichia coli/genetics; Humans; Models, Molecular}},
  language     = {{eng}},
  month        = {{10}},
  number       = {{Pt 10}},
  pages        = {{895--903}},
  publisher    = {{John Wiley & Sons Inc.}},
  series       = {{Acta Crystallographica Section D: Structural Biology}},
  title        = {{Structural comparison of protiated, H/D-exchanged and deuterated human carbonic anhydrase IX}},
  url          = {{http://dx.doi.org/10.1107/S2059798319010027}},
  doi          = {{10.1107/S2059798319010027}},
  volume       = {{75}},
  year         = {{2019}},
}