MOLECULAR-STRUCTURE OF KANAMYCIN NUCLEOTIDYLTRANSFERASE DETERMINED TO3.0-ANGSTROM RESOLUTION

Kanamycin nucleotidyltransferase, as originally isolated from Staphylo coccus aureus, inactivates the antibiotic kanamycin by catalyzing the transfer of a nucleotidyl group from nucleoside triphosphates such as ATP to the 4'-hydroxyl group of the aminoglycoside. The molecular stru cture of the enzyme described here was determined by X-ray crystallogr aphic analysis to are solution of 3.0 angstrom. Crystals employed in t he investigation belonged to the space group P4(3)2(1)2 with unit cell dimensions of a = b = 78.9 angstrom and c = 219.2 angstrom. An electr on density map phased with seven heavy-atom derivatives revealed that the molecules packed in the crystalline lattice as dimers exhibiting l ocal 2-fold rotation axes. Subsequent symmetry averaging and solvent f lattening improved the quality of the electron density such that it wa s possible to completely trace the 253 amino acid polypeptide chain. E ach monomer is divided into two distinct structural domains: the N-ter minal motif composed of residues Met 1-Glu 127 and the C-terminal half delineated by residues Ala 128-Phe 253. The N-terminal region is char acterized by a five-stranded mixed beta-pleated sheet whereas the C-te rminal domain contains five alpha-helices, four of which form an up-an d-down alpha-helical bundle very similar to that observed in cytochrom e c'. The two subunits wrap about one another to form an ellipsoid wit h a pronounced cleft that could easily accommodate the various aminogl ycosides known to bind to the enzyme.