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Direct contacts between conserved motifs of different subunits provide major contribution to active site organization in human and mycobacterial dUTPases
Author(s) -
Takács Enikő,
Nagy Gergely,
Leveles Ibolya,
Harmat Veronika,
Lopata Anna,
Tóth Judit,
Vértessy Beáta G.
Publication year - 2010
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/j.febslet.2010.05.018
Subject(s) - active site , conserved sequence , mutant , protein subunit , binding site , sequence alignment , protein structure , biology , enzyme , chemistry , biochemistry , genetics , peptide sequence , gene
dUTP pyrophosphatases (dUTPases) are essential for genome integrity. Recent results allowed characterization of the role of conserved residues. Here we analyzed the Asp/Asn mutation within conserved Motif I of human and mycobacterial dUTPases, wherein the Asp residue was previously implicated in Mg 2+ ‐coordination. Our results on transient/steady‐state kinetics, ligand binding and a 1.80 Å resolution structure of the mutant mycobacterial enzyme, in comparison with wild type and C‐terminally truncated structures, argue that this residue has a major role in providing intra‐ and intersubunit contacts, but is not essential for Mg 2+ accommodation. We conclude that in addition to the role of conserved motifs in substrate accommodation, direct subunit interaction between protein atoms of active site residues from different conserved motifs are crucial for enzyme function.