Promiscuity in biosynthetic enzymes: structural basis for antibiotic diversity

Abstract

Aromatic polyketide antibiotics are of particularmedical importance, because some of the clinically most potent anti-tumor drugs are recruited from this class of compounds. A striking feature of many of the enzymes in aromatic polyketide biosynthesis is their relative broad substrate specificity, i.e. they accept intermediates from several related biosynthetic pathways as substrates. This feature is the basis of present-day combinatorial biosynthesis efforts to generate antibiotic diversity, where genes from the biosynthetic cluster of one species are transferred to another species, the hybrid antibiotic approach. Our laboratory is engaged in a small-scale structural genomics project aiming at the structural and functional characterisation of enzymes in the biosynthesis of aromatic polyketide antibiotics, in particular anthracyclines. Out of approximately 50 target genes, we have at present cloned twenty-two genes and produced sixteen enzymes in soluble form. Twelve of these have been crystallized and the crystal structures of ten enzymes have been determined so far. Several of these enzymes show novel mechanisms, for instance the polyketide cyclase SnoaL or the S-adenosyl-L-methionine dependent hydroxylase RdmB. The latter case is the first observation of S-adenosyl L-methionine as a cofactor in an enzymatic hydroxylation reaction. These studies have also provided insights into the structural basis of substrate recognition and specificity of these enzymes. Binding and recognition of the polyketide substrates is dominated by hydrophobic interactions. Specificity is controlled by the shape of the binding pocket rather than through specific hydrogen bonds. Re-design of the substrate binding pockets could possibly expand the substrate spectrum of these enzymes, and thus provide a larger repertoire of biosynthetic enzymes for combinatorial biosynthesis. This approach could therefore potentially facilitate the development of novel aromatic polyketide antibiotics with improved toxicity profiles. m11.o03