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Crystal Structure of dUTPase from Equine Infectious Anaemia Virus; Active Site Metal Binding in a Substrate Analogue Complex

Dauter, Zbigniew; Persson, Rebecca; Rosengren, Anna Maria; Nyman, Per-Olof LU ; Wilson, Keith S and Cedergren-Zeppezauer, Eila S (1999) In Journal of Molecular Biology 285(2). p.655-673
Abstract
The X-ray structures of dUTPase from equine infectious anaemia virus (EIAV) in unliganded and complexed forms have been determined to 1.9 and 2.0 A resolution, respectively. The structures were solved by molecular replacement using Escherichia coli dUTPase as search model. The exploitation of a relatively novel refinement approach for the initial model, combining maximum likelihood refinement with stereochemically unrestrained updating of the model, proved to be of crucial importance and should be of general relevance.EIAV dUTPase is a homotrimer where each subunit folds into a twisted antiparallel @b-barrel with the N and C-terminal portions interacting with adjacent subunits. The C-terminal 14 and 17 amino acid residues are disordered in... (More)
The X-ray structures of dUTPase from equine infectious anaemia virus (EIAV) in unliganded and complexed forms have been determined to 1.9 and 2.0 A resolution, respectively. The structures were solved by molecular replacement using Escherichia coli dUTPase as search model. The exploitation of a relatively novel refinement approach for the initial model, combining maximum likelihood refinement with stereochemically unrestrained updating of the model, proved to be of crucial importance and should be of general relevance.EIAV dUTPase is a homotrimer where each subunit folds into a twisted antiparallel @b-barrel with the N and C-terminal portions interacting with adjacent subunits. The C-terminal 14 and 17 amino acid residues are disordered in the crystal structure of the unliganded and complexed enzyme, respectively. Interactions along the 3-fold axis include a water-containing volume (size 207 A3) which has no contact with bulk solvent.It has earlier been shown that a divalent metal ion is essential for catalysis. For the first time, a putative binding site for such a metal ion, in this case Sr2+, is established. The positions of the inhibitor (the non-hydrolysable substrate analogue dUDP) and the metal ion in the complex are consistent with the location of the active centre established for trimeric dUTPase structures, in which subunit interfaces form three surface clefts lined with evolutionary conserved residues. However, a detailed comparison of the active sites of the EIAV and E. coli enzymes reveals some structural differences. The viral enzyme undergoes a small conformational change in the uracil-binding @b-hairpin structure upon dUDP binding not observed in the other known dUTPase structures. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
X-ray structure, dUTPase, metal-inhibitor complex, equine infectious anaemia virus, automated refinement procedure
in
Journal of Molecular Biology
volume
285
issue
2
pages
655 - 673
publisher
Elsevier
external identifiers
  • scopus:0033555756
ISSN
1089-8638
DOI
10.1006/jmbi.1998.2332
language
English
LU publication?
yes
id
1f8b4396-c853-4461-9f57-c09be6172b9e (old id 125268)
date added to LUP
2007-07-04 16:07:05
date last changed
2017-04-09 04:11:52
@article{1f8b4396-c853-4461-9f57-c09be6172b9e,
  abstract     = {The X-ray structures of dUTPase from equine infectious anaemia virus (EIAV) in unliganded and complexed forms have been determined to 1.9 and 2.0 A resolution, respectively. The structures were solved by molecular replacement using Escherichia coli dUTPase as search model. The exploitation of a relatively novel refinement approach for the initial model, combining maximum likelihood refinement with stereochemically unrestrained updating of the model, proved to be of crucial importance and should be of general relevance.EIAV dUTPase is a homotrimer where each subunit folds into a twisted antiparallel @b-barrel with the N and C-terminal portions interacting with adjacent subunits. The C-terminal 14 and 17 amino acid residues are disordered in the crystal structure of the unliganded and complexed enzyme, respectively. Interactions along the 3-fold axis include a water-containing volume (size 207 A3) which has no contact with bulk solvent.It has earlier been shown that a divalent metal ion is essential for catalysis. For the first time, a putative binding site for such a metal ion, in this case Sr2+, is established. The positions of the inhibitor (the non-hydrolysable substrate analogue dUDP) and the metal ion in the complex are consistent with the location of the active centre established for trimeric dUTPase structures, in which subunit interfaces form three surface clefts lined with evolutionary conserved residues. However, a detailed comparison of the active sites of the EIAV and E. coli enzymes reveals some structural differences. The viral enzyme undergoes a small conformational change in the uracil-binding @b-hairpin structure upon dUDP binding not observed in the other known dUTPase structures.},
  author       = {Dauter, Zbigniew and Persson, Rebecca and Rosengren, Anna Maria and Nyman, Per-Olof and Wilson, Keith S and Cedergren-Zeppezauer, Eila S},
  issn         = {1089-8638},
  keyword      = {X-ray structure,dUTPase,metal-inhibitor complex,equine infectious anaemia virus,automated refinement procedure},
  language     = {eng},
  number       = {2},
  pages        = {655--673},
  publisher    = {Elsevier},
  series       = {Journal of Molecular Biology},
  title        = {Crystal Structure of dUTPase from Equine Infectious Anaemia Virus; Active Site Metal Binding in a Substrate Analogue Complex},
  url          = {http://dx.doi.org/10.1006/jmbi.1998.2332},
  volume       = {285},
  year         = {1999},
}