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Crystal structure of human inosine triphosphatase : Substrate binding and implication of the inosine triphosphatase deficiency mutation P32T

Stenmark, Pål LU orcid ; Kursula, Petri ; Flodin, Susanne ; Gräslund, Susanne ; Landry, Robert ; Nordlund, Pär and Schüler, Herwig LU orcid (2007) In The Journal of biological chemistry 282(5). p.3182-3187
Abstract

Inosine triphosphatase (ITPA) is a ubiquitous key regulator of cellular non-canonical nucleotide levels. It breaks down inosine and xanthine nucleotides generated by deamination of purine bases. Its enzymatic action prevents accumulation of ITP and reduces the risk of incorporation of potentially mutagenic inosine nucleotides into nucleic acids. Here we describe the crystal structure of human ITPA in complex with its prime substrate ITP, as well as the apoenzyme at 2.8 and 1.1A, respectively. These structures show for the first time the site of substrate and Mg2+ coordination as well as the conformational changes accompanying substrate binding in this class of enzymes. Enzyme substrate interactions induce an extensive closure of the... (More)

Inosine triphosphatase (ITPA) is a ubiquitous key regulator of cellular non-canonical nucleotide levels. It breaks down inosine and xanthine nucleotides generated by deamination of purine bases. Its enzymatic action prevents accumulation of ITP and reduces the risk of incorporation of potentially mutagenic inosine nucleotides into nucleic acids. Here we describe the crystal structure of human ITPA in complex with its prime substrate ITP, as well as the apoenzyme at 2.8 and 1.1A, respectively. These structures show for the first time the site of substrate and Mg2+ coordination as well as the conformational changes accompanying substrate binding in this class of enzymes. Enzyme substrate interactions induce an extensive closure of the nucleotide binding grove, resulting in tight interactions with the base that explain the high substrate specificity of ITPA for inosine and xanthine over the canonical nucleotides. One of the dimer contact sites is made up by a loop that is involved in coordinating the metal ion in the active site. We predict that the ITPA deficiency mutation P32T leads to a shift of this loop that results in a disturbed affinity for nucleotides and/or a reduced catalytic activity in both monomers of the physiological dimer.

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author
; ; ; ; ; and
publishing date
type
Contribution to journal
publication status
published
keywords
Amino Acid Substitution, Binding Sites, Cloning, Molecular, Crystallization, Crystallography, X-Ray, DNA, Complementary, Humans, Models, Molecular, Mutation, Protein Conformation, Protein Structure, Secondary, Pyrophosphatases/chemistry, Recombinant Proteins/chemistry, Inosine Triphosphatase
in
The Journal of biological chemistry
volume
282
issue
5
pages
6 pages
publisher
American Society for Biochemistry and Molecular Biology
external identifiers
  • pmid:17138556
  • scopus:34047247010
ISSN
0021-9258
DOI
10.1074/jbc.M609838200
language
English
LU publication?
no
id
6509acc3-5cc3-4942-89ae-d6b7722f7677
date added to LUP
2024-11-21 18:05:04
date last changed
2025-01-09 03:13:02
@article{6509acc3-5cc3-4942-89ae-d6b7722f7677,
  abstract     = {{<p>Inosine triphosphatase (ITPA) is a ubiquitous key regulator of cellular non-canonical nucleotide levels. It breaks down inosine and xanthine nucleotides generated by deamination of purine bases. Its enzymatic action prevents accumulation of ITP and reduces the risk of incorporation of potentially mutagenic inosine nucleotides into nucleic acids. Here we describe the crystal structure of human ITPA in complex with its prime substrate ITP, as well as the apoenzyme at 2.8 and 1.1A, respectively. These structures show for the first time the site of substrate and Mg2+ coordination as well as the conformational changes accompanying substrate binding in this class of enzymes. Enzyme substrate interactions induce an extensive closure of the nucleotide binding grove, resulting in tight interactions with the base that explain the high substrate specificity of ITPA for inosine and xanthine over the canonical nucleotides. One of the dimer contact sites is made up by a loop that is involved in coordinating the metal ion in the active site. We predict that the ITPA deficiency mutation P32T leads to a shift of this loop that results in a disturbed affinity for nucleotides and/or a reduced catalytic activity in both monomers of the physiological dimer.</p>}},
  author       = {{Stenmark, Pål and Kursula, Petri and Flodin, Susanne and Gräslund, Susanne and Landry, Robert and Nordlund, Pär and Schüler, Herwig}},
  issn         = {{0021-9258}},
  keywords     = {{Amino Acid Substitution; Binding Sites; Cloning, Molecular; Crystallization; Crystallography, X-Ray; DNA, Complementary; Humans; Models, Molecular; Mutation; Protein Conformation; Protein Structure, Secondary; Pyrophosphatases/chemistry; Recombinant Proteins/chemistry; Inosine Triphosphatase}},
  language     = {{eng}},
  month        = {{02}},
  number       = {{5}},
  pages        = {{3182--3187}},
  publisher    = {{American Society for Biochemistry and Molecular Biology}},
  series       = {{The Journal of biological chemistry}},
  title        = {{Crystal structure of human inosine triphosphatase : Substrate binding and implication of the inosine triphosphatase deficiency mutation P32T}},
  url          = {{http://dx.doi.org/10.1074/jbc.M609838200}},
  doi          = {{10.1074/jbc.M609838200}},
  volume       = {{282}},
  year         = {{2007}},
}