Crystal structure of the glycosyltransferase SnogD from the biosynthetic pathway of nogalamycin in Streptomyces nogalater

Abstract

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. DatabaseThe atomic coordinates and structure factors have been deposited with the RCSB Protein Data Bank under accession numbers 4AMB, 4AMG and 4AN4 Structured digital abstract l snogD and snogD bind by x-ray crystallography (View Interaction: 1, 2)

A pharmaceutical model for the molecular evolution of microbial natural products

“…The canonical reaction of the anthracycline fourth ring polyketide cyclases such as SnoaL and AknH is to catalyze aldol condensations that determine the stereochemical configuration at C9, but interestingly SnoaL2 has also been observed to harbor minor ancestral cyclization activity . After attachment of l ‐rhodosamine by the glycosyltransferase SnogD , the subtype epitomizing C2–C5´´ bond is formed by the 2OG and nonhaem iron‐dependent oxygenase SnoK . However, a gene duplication event has led to the appearance of C4´´ epimerase SnoN from SnoK .…”

Advances in understanding glycosyltransferases from a structural perspective

“…Nogalamine is attached to the polyketide via both an O -glycosidic bond and C—C linkage. The structure of SnogD, in complex with nucleotide, supports the mechanism for the formation of the O -glycosidic bond, but it is less clear how SnogD catalyses the synthesis of a C—C bond, which may in fact be generated by a different enzyme [ 40 ]. The GTs capable of transferring glucose to anthocyanidins [ 41 ] and of transferring olivose to a tetracycline-like scaffold via the formation of a C—C bond [ 42 ] have also been investigated.…”

Identification of the incednine biosynthetic gene cluster: characterization of novel β-glutamate-β-decarboxylase IdnL3

“…IdnS9 and IdnS15 lack many internal protein sequences conserved with the other known glycosyltransferases, as well as a crucial histidine residue that is critical for the deprotonation of the hydroxy group of the glycosyl acceptor molecules (Supplementary Figure S4). 38,39 Therefore, IdnS9 and IdnS15 appear to be inactive proteins, and IdnS4 and IdnS14 are predicted to be involved in the two glycosylation events, the attachment of N-methylxylosamine and N-demethylforosamine, although functional characterization of these glycosyltransferases is necessary to elucidate the precise biosynthetic role.…”