title: Mechanistic studies of dUTPases.
abstract: The essential enzyme dUTPase is responsible for preventive DNA repair via exclusion of uracil. Lack or inhibition of the enzyme induces thymine-less cell death in cells performing active DNA synthesis, serving therefore as an important chemotherapeutic target. In the present work, employing differential circular dichroism spectroscopy, we show that D. mel. dUTPase, a recently described eukaryotic model, has a similar affinity of binding towards alpha,beta-imino-dUTP as compared to the prokaryotic E. coli enzyme. However, in contrast to the prokaryotic dUTPase, the nucleotide exerts significant protection against tryptic digestion at a specific tryptic site 20 A far from the active site in the fly enzyme. This result indicates that binding of the nucleotide in the active site induces an allosteric conformational change within the central threefold channel of the homotrimer exclusively in the eukaryotic enzyme. Nucleotide binding induced allosterism in the D. mel. dUTPase, but not in the E. coli enzyme, might be associated with the altered hydropathy of subunit interfaces in these two proteins.
authors.affiliation: Inst. Enzymol., Hung. Acad. Sci., Budapest, Hungary.
authors.forename: J
authors.initials: J
authors.lastname: Kovári
authors.forename: T
authors.initials: T
authors.lastname: Imre
authors.forename: P
authors.initials: P
authors.lastname: Szabó
authors.forename: B G
authors.initials: BG
authors.lastname: Vértessy
mesh_terms: D000495:Allosteric Site; D000818:Animals; D001665:Binding Sites; D002942:Circular Dichroism; D004247:DNA / Q000737:chemistry*; D004260:DNA Repair; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004331:Drosophila melanogaster; D004926:Escherichia coli / Q000201:enzymology / Q000378:metabolism; D008958:Models, Molecular; D011487:Protein Conformation; D011755:Pyrophosphatases / Q000737:chemistry* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D014357:Trypsin / Q000737:chemistry / Q000494:pharmacology; D014498:Uracil / Q000737:chemistry
chemical_list: D011994:Recombinant Proteins; D014498:Uracil; D004247:DNA; D014357:Trypsin; D011755:Pyrophosphatases; C020020:dUTP pyrophosphatase