Crystal structure of the glycosyltransferase SnogD from the biosynthetic pathway of nogalamycin in Streptomyces nogalater.
The glycosyltransferase SnogD from Streptomyces nogalater transfers a nogalamine moiety to the metabolic intermediate 3',4'-demethoxynogalose-1-hydroxynogalamycinone during the final steps of biosynthesis of the aromatic polyketide nogalamycin. The crystal structure of recombinant SnogD, as an apo-enzyme and with a bound nucleotide, 2-deoxyuridine-5'-diphosphate, was determined to 2.6 Å resolution. Reductive methylation of SnogD was crucial for reproducible preparation of diffraction quality crystals due to creation of an additional intermolecular salt bridge between methylated lysine residue Lys384 and Glu374* from an adjacent molecule in the crystal lattice. SnogD is a dimer both in solution and in the crystal, and the enzyme subunit displays a fold characteristic of the GT-B family of glycosyltransferases. Binding of the nucleotide is associated with rearrangement of two active-site loops. Site-directed mutagenesis shows that two active-site histidine residues, His25 and His301, are critical for the glycosyltransferase activities of SnogD both in vivo and in vitro. The crystal structures and the functional data are consistent with a role for His301 in binding of the diphosphate group of the sugar donor substrate, and a function of His25 as a catalytic base in the glycosyl transfer reaction.
Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.
Magnus
M
Claesson
Vilja
V
Siitonen
Doreen
D
Dobritzsch
Mikko
M
Metsä-Ketelä
Gunter
G
Schneider
D000903:Antibiotics, Antineoplastic / Q000096:biosynthesis*; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D016695:Glycosyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D009621:Nogalamycin / Q000096:biosynthesis*; D011487:Protein Conformation; D013302:Streptomyces / Q000378:metabolism*
D000903:Antibiotics, Antineoplastic; D016695:Glycosyltransferases; D009621:Nogalamycin