Codon-anticodon interaction at the ribosomal E site.
The question of whether or not the tRNA at the third ribosomal binding site specific for deacylated tRNA (E site) undergoes codon-anticodon interaction was analyzed as follows. Poly(U)-programmed ribosomes each carrying two [14C]tRNAPhe molecules were subjected to a chasing experiment using various tRNA species. At 0 degree C Ac[3H]Phe-tRNAPhe did not trigger any chasing whereas deacylated cognate tRNAPhe provoked a strong effect; non-cognate tRNALys was totally ineffective. This indicates that the second [14C]tRNAPhe cannot be present at the A site but rather at the E site (confirming previous observations). In the presence of poly(U) or poly(A) ribosomes bound the cognate tRNA practically exclusively as second deacylated tRNA, i.e. [14C]tRNAPhe and [14C]tRNALys, respectively. Thus, the second deacylated tRNA binds in a codon-dependent manner. [14C]tRNALys at the P site and Ac[3H]Lys-tRNALys at the A site of poly(A)-primed ribosomes were translocated to the E and P sites, respectively, by means of elongation factor G. The E site-bound [14C]tRNALys could be significantly chased by cognate tRNALys but not by non-cognate tRNAPhe, indicating the coded nature of the E site binding. Additional evidence is presented that the ribosome accommodates two adjacent codon-anticodon interactions at either A and P or P and E sites.
D000215:Acylation; D000926:Anticodon / Q000378:metabolism*; D001665:Binding Sites; D001667:Binding, Competitive; D003062:Codon / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D020653:Peptide Elongation Factor G; D010445:Peptide Elongation Factors / Q000378:metabolism; D011061:Poly A / Q000494:pharmacology; D011072:Poly U / Q000494:pharmacology; D012329:RNA, Bacterial / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism*; D012343:RNA, Transfer / Q000378:metabolism*; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism*
D000926:Anticodon; D003062:Codon; D020653:Peptide Elongation Factor G; D010445:Peptide Elongation Factors; D012329:RNA, Bacterial; D012333:RNA, Messenger; D012346:RNA, Transfer, Amino Acyl; D011061:Poly A; D011072:Poly U; D012343:RNA, Transfer