23S RIBOSOMAL-RNA POSITIONS ESSENTIAL FOR TRANSFER-RNA BINDING IN RIBOSOMAL FUNCTIONAL SITES

rRNA plays an important role in function of peptidyl transferase, the catalytic center of the ribosome responsible for the peptide bond form ation, Proper placement of the peptidyl transferase substrates, peptid yl-tRNA and aminoacyl-tRNA, is essential for catalysis of the transpep tidation reaction and protein synthesis, In this report, we define a s mall set of rRNA nucleotides that are most likely directly involved in binding of tRNA in the functional sites of the large ribosomal subuni t, By binding biotinylated tRNA substrates to randomly modified large ribosomal subunits from Escherichia coli and capturing resulting compl exes on the avidin resin, we identified four nucleotides in the large ribosomal subunit rRNA (positions G2252, A2451, U2506, and U2585) whos e modifications prevent binding of a peptidyl-tRNA analog in the P sit e and one residue (U2555) whose modification interferes with transfer of peptidyl moiety to puromycin, These nucleotides represent a subset of positions protected by tRNA analogs from chemical modification and significantly narrow the number of 23S rRNA nucleotides that may be di rectly involved in tRNA binding in the ribosomal functional sites.