ABSTRACT
The best-known orthosomycin compounds are evernimicin that was developed for use in humans but never made it to the market and avilamycin that is used as a growth promoter. The orthosomycin antibiotics target a site on the 50S ribosomal subunit that includes protein LI 6 and 23S rRNA helices 89 and 91 and is close to both the elbow of tRNA positioned in the A-site47 (Fig. 1) and the initiation factor 2 binding site.48 Mutations in ribosomal protein L16 and in domain V of 23S rRNA (at positions 2469-72,2479-80,2535-6) confer resistance to orthosomycins (sum marised in ref. 46). The EmtA methyltransferase acting at 23S rRNA position G2470 was found in Enterococcusfaecium from animal sources on the basis o f its resistance to the growth promoter avilamycin and it also confers resistance to evernimicin (Table l ).49 It was shown that purified EmtA methylated 50S subunits from an evernimicin-sensitive strain 30-fold more efficiendy than those from a resistant strain49 indicating that the entire 50S subunit can act as a substrate for this methyltransferase. Soon after this discovery, four genes in the natural producer o f avilamycin, the actinomycetes Streptomyces viridochromogenes Tit57, were found to be resistance determinants.50 Two o f the genes are similar to an ATP binding cassette transporter system that probably exports avilamycin across the cell membrane, whereas the other two genes, aviRa and aviRb, encode rRNA methyltransferases. The modifications were later identified as a 2'-0 -ribose methylation at U2479 conferred by AviRb and abase methylation at G2535 conferred by AviRa (Table l ).51 Both of these sites are in the same RNA region on the 50S subunit as all the resistance mutations. AviRb confers a higher level o f resistance than AviRa, but both are necessary for self-protection in the avilamy cin producer and they are thought to act together like the tylosin resistance methyltransferases described above. The crystal structure o f the AviRa methyltransferase has been determined by X-ray diffraction at 1.5 A resolution.52 Its overall fold is similar to that o f most methyltransferases, but it contains two additional helices as a specific feature. Guided by the target, the enzyme was docked to the cognate ribosomal surface, where it fit well into a deep cleft without contacting any ribosomal proteins. A putative-binding site for the SAM cofactor was derived from homologous structures, but because the transferred methyl group is in a pocket beneath the enzyme surface, the targeted guanine base has to flip out for methylation.52
