Crystal structures of nucleoside 2-deoxyribosyltransferase in native and ligand-bound forms reveal architecture of the active site.
BACKGROUND
Nucleoside 2-deoxyribosyltransferase plays an important role in the salvage pathway of nucleotide metabolism in certain organisms, catalyzing the cleavage of beta-2'-deoxyribonucleosides and the subsequent transfer of the deoxyribosyl moiety to an acceptor purine or pyrimidine base. The kinetics describe a ping-pong-bi-bi pathway involving the formation of a covalent enzyme-deoxyribose intermediate. The enzyme is produced by a limited number of microorganisms and its functions have been exploited in its use as a biocatalyst to synthesize nucleoside analogs of therapeutic interest.


RESULTS
We describe the crystal structure of the enzyme with and without bound ligand. The native structure was solved by the single isomorphous replacement with anomalous scattering method (SIRAS) and refined to 2.5 A resolution resulting in a crystallographic R factor of 16.6%. The enzyme comprises a single domain that belongs to the general class of doubly-wound alpha/beta proteins; it also exhibits a unique nucleoside-binding motif. X-ray analysis of enzyme-purine and enzyme-pyrimidine complexes presented here reveals that the active site lies in a cleft formed by the edge of the beta sheet and two alpha helices and contains side chains from two subunits.


CONCLUSIONS
These results indicate residues that may be important in substrate binding and catalysis and thus may serve as a framework for elucidating the mechanism of enzyme activity. In particular, the proposed nucleophile, Glu98, lies in the nucleoside-binding pocket at an appropriate position for nucleophilic attack. A comparison of the enzyme interactions with both a purine and pyrimidine ligand provides some insight into the structural basis for enzyme specificity.
Armstrong
S R
SR
Section of Biochemistry, Cell and Molecular Biology, Cornell University, Ithaca, NY 14853, USA.
Cook
W J
WJ
Short
S A
SA
Ealick
S E
SE
D001665:Binding Sites; D018360:Crystallography, X-Ray; D007778:Lactobacillus / Q000201:enzymology; D010430:Pentosyltransferases / Q000737:chemistry*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011687:Purines / Q000737:chemistry / Q000378:metabolism; D011743:Pyrimidines / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity
D011687:Purines; D011743:Pyrimidines; D010430:Pentosyltransferases; C021888:nucleoside deoxyribosyltransferase