Affinity labelling of Escherichia coli ribosomes with a benzylidene derivative of AUGU sub(6) within initiation and pretranslocational complexes

Affinity labelling of E. coli ribosomes with the 2',3'-O-(4-(N-2-chloroethyl)-N-methylamino)benzylidene derivative of AUGU sub(6) was studied within the initiation complex (complex I) obtained by using fMet-tRNA sub(f) super(M)eta)nd initiation factors and within the pretranslocational complex (complex II) obtained by treatment of complex I with the ternary complex Phe-tRNA super(Phe) multiplied by GTP multiplied by EF-Tu. Both proteins and rRNA of 30 S as well as 50 S subunits were found to be labelled. Sets of proteins labelled within complexes I and II differ considerably. Within complex II, proteins S13 and L10 were labelled preferentially. On the other hand, within complex I, multiple modification is observed (proteins S4, S12, S13, S14, S15, S18, S19, S20/L26 were found to be alkylated) despite the single fixation of a template in the ribosome by interaction of the AUG codon with fMet-tRNA sub(f) super(M)et )