Catalytic site conformations in human PNP by 19F-NMR and crystallography

Javier Suarez, Antti M. Haapalainen, Sean M. Cahill, Meng Chiao Ho, Funing Yan, Steven C. Almo, Vern L. Schramm

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

Purine nucleoside phosphorylase (PNP) is a target for leukemia, gout, and autoimmune disorders. Dynamic motion of catalytic site loops has been implicated in catalysis, but experimental evidence was lacking. We replaced catalytic site groups His257 or His64 with 6-fluoro-tryptophan (6FW) as site-specific NMR probes. Conformational adjustments in the 6FW-His257-helical and His64-6FW-loop regions were characterized in PNP phosphate-bound enzyme and in complexes with catalytic site ligands, including transition state analogs. Chemical shift and line-shape changes associated with these complexes revealed dynamic coexistence of several conformational states in these regions in phosphate-bound enzyme and altered or single conformations in other complexes. These conformations were also characterized by X-ray crystallography. Specific 19F-Trp labels and X-ray crystallography provide multidimensional characterization of conformational states for free, catalytic, and inhibited complexes of human PNP.

Original languageEnglish (US)
Pages (from-to)212-222
Number of pages11
JournalChemistry and Biology
Volume20
Issue number2
DOIs
StatePublished - Feb 21 2013

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Purine-Nucleoside Phosphorylase
Crystallography
Conformations
Catalytic Domain
X ray crystallography
X Ray Crystallography
Nuclear magnetic resonance
Phosphates
Gout
Chemical shift
Enzymes
Catalysis
Tryptophan
Labels
Leukemia
Ligands

ASJC Scopus subject areas

  • Biochemistry
  • Drug Discovery
  • Molecular Biology
  • Clinical Biochemistry
  • Molecular Medicine
  • Pharmacology

Cite this

Catalytic site conformations in human PNP by 19F-NMR and crystallography. / Suarez, Javier; Haapalainen, Antti M.; Cahill, Sean M.; Ho, Meng Chiao; Yan, Funing; Almo, Steven C.; Schramm, Vern L.

In: Chemistry and Biology, Vol. 20, No. 2, 21.02.2013, p. 212-222.

Research output: Contribution to journalArticle

Suarez, Javier ; Haapalainen, Antti M. ; Cahill, Sean M. ; Ho, Meng Chiao ; Yan, Funing ; Almo, Steven C. ; Schramm, Vern L. / Catalytic site conformations in human PNP by 19F-NMR and crystallography. In: Chemistry and Biology. 2013 ; Vol. 20, No. 2. pp. 212-222.
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