Streptomycin binding sites

Upon irradiation (incandescent lamp, 400 W) of the reaction mixtures of the streptomycin analog with 30 S or 50 S ribosomal subparticles taken separately, the analog covalently binds predominantly to the 30 S subparticle. There is no covalent binding without irradiation. Therefore, we have concluded that the compound is an active photoafiinity analog of streptomycin. An analysis of components labeled with this analog permitted identification of the proteins of the 30 S ribosomal subparticle located in or near the streptomycin-binding site, i.e., proteins S 7 and S 14. 

Alteration in binding site Streptomycin Protein S12 component of 30S ribosomal subunit determines sensitivity or resistance... 

Alterations in the binding site of ribosomal protein SI2, accounts for high-level insusceptibility to streptomycin in E. coli and S. aureus [166] whereas mutations in RNA polymerase are associated with decreased sensitivity to rifampicin [167], Other chromosomal mutational examples involving insensitive target enzymes occur with modified dihydrofolate reductases and dihydroperoate synthetases [168], Chromosomal mutations in E. coli cause overproduction of the target enzyme for trimethoprim (34) activity [169]. Mutation of target enzyme in Strep, pneumoniae provides lower sul-... 

Resistance to aminoglycosides arises from several mechanisms, the most important being the production of enzymes (plasmid controlled) that inactivate the drug by acetylation, phosphorylation or adenylation. Other mechanisms are the alterations of the envelope to prevent drug access and alteration of the binding site on the 30S subunit so that the drug does not bind (streptomycin only). 

Tests of Function. Modified antibiotics were compared to the natural compounds in reversible binding to ribosomes and in inhibitory or other effects on ribosome function. In one case, the examination also included the in vivo effects of an antibiotic analog. In in vitro experiments, the interference by irreversible binding was prevented or minimized by performing the tests under conditions unfavorable for covalent reaction, e.g., low temperatures or suboptimal pH. Reversible binding studies were carried out with the clhoramphenicol and streptomycin derivatives and included the subunit localization of binding sites, estimation of the... 

The binding sites of streptomycin and of puromycin on bacterial ribosomes have been probed with chemically reactive derivatives of both... 

Puromycin binds weakly and relatively unspecifically to ribosomes. Its use as an affinity probe is limited. Not unexpectedly, reactions of iV-iodoacetylpuromycin with ribosomes exhibit similar characteristics. On the other hand, a high binding constant does not necessarily guarantee a good affinity probe as exemplified by the streptomycin affinity labels. Vice versa, a compound with a weak binding constant might still be a superb affinity probe, since the outcome of experiments with halo-acetyl compounds depends so much on the presence of a properly oriented reactive amino acid side chain in the substrate binding site. This, of course, is a matter of chance. ... 

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