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The Crystal Structure of Streptococcus pyogenes Uridine Phosphorylase Reveals a Distinct Subfamily of Nucleoside Phosphorylases

In English

Författare

  • Timothy H. Tran
  • Stig Christoffersen
  • Paula W. Allan
  • William B. Parker
  • Jure Piskur
  • I. Serra
  • M. Terreni
  • Steven E. Ealick

Summary, in English

Uridine phosphorylase (UP), a key enzyme in the pyrimidine salvage pathway catalyzes the reversible phosphorolysis of uridine or 2'-deoxyuridine to uracil and ribose 1-phosphate or 2'-deoxyribose 1-phosphate. This enzyme belongs to the nucleoside phosphorylase I superfamily whose members show diverse specificity for nucleoside substrates. Phylogenetic analysis shows Streptococcus pyogenes uridine phosphorylase (SpUP) is found in a distinct branch of the pyrimidine subfamily of nucleoside phosphorylases. To further characterize SpUP, we determined the crystal structure in complex with the products, ribose I-phosphate and uracil, at 1.8 angstrom resolution. Like Escherichia coli UP (EcUP), the biological unit of SpUP is a hexamer with an alpha/beta monomeric fold. A novel feature of the active site is the presence of His169, which structurally aligns with Arg168 of the EcUP structure. A second active site residue, Lys162, is not present in previously determined UP structures and interacts with O2 of uracil. Biochemical studies of wild-type SpUTP showed that its substrate specificity is similar to that of EcUP, while EcUP is similar to 7-fold more efficient than SpUP. Biochemical studies of SpUP mutants showed that mutations of His 169 reduced activity, while mutation of Lys162 abolished all activity, suggesting that the negative charge in the transition state resides mostly on uracil O2. This is in contrast to EcUP for which transition state stabilization occurs mostly at O4.

Publiceringsår

2011

Språk

Engelska

Sidor

6549-6558

Publikation/Tidskrift/Serie

Biochemistry

Volym

50

Issue

30

Dokumenttyp

Artikel i tidskrift

Förlag

The American Chemical Society (ACS)

Ämne

  • Biochemistry and Molecular Biology

Status

Published

ISBN/ISSN/Övrigt

  • ISSN: 0006-2960