CONTACT PATTERNS OF RNA LIGANDS WITH THE RIBOSOME IN DEFINED FUNCTIONAL-STATES AS DETERMINED BY PROTECTION AGAINST CLEAVAGE AT PHOSPHOROTHIOATED RESIDUES - A REVIEW

A few years ago a new footprinting method was introduced which identif ies contacts at the phosphate groups of RNAs, The method exploits the potential of iodine (I-2) to cleave at phosphorothioated positions of RNAs as long as no other components prevent access of the iodine molec ule to the corresponding phosphorothioate group. Here we give a survey of recent experiments where the contact patterns of mRNA and tRNAs wi thin active ribosomes have been explored. The initiation state of ribo somes as well as the two elongation states, viz. the pre- and the post -translocational states, have been analyzed. Only weak protections wer e found for the mRNA. In the elongation complexes the phosphate group two positions upstream of the decoding codons was protected, whereas i n the initiation complex phosphate groups in the Shine-Dalgarno sequen ce were also protected. No protection could be seen downstream of the decoding codons. In contrast, numerous phosphate positions were protec ted in tRNAs bound to the ribosome. The tRNA protection patterns were highly differentiated and were strikingly different for the two tRNAs simultaneously bound to the ribosome. The protection pattern of an ind ividual tRNA remained almost constant upon translocation. The results suggest that the ribosome might hold the tRNAs by a whole set of direc t contacts, whereas the mRNA is mainly fixed at the two decoding codon s and by the tRNAs via codon-anticodon interaction. A hypothesis about a possible mechanism of the translocation process is discussed.