AFFINITY LABELING OF HUMAN PLACENTA-80S R IBOSOMES WITH MESSENGER-RNAANALOGS-DERIVATIVES OF OLIGOURIDYLATES BEARING AN ALKYLATING GROUP AT5'-END

Affinity labelling of 80S ribosomes from human placenta with 4-(N-meth ylamino-N-2-chloroethyl) benzylmethylphosphoramide derivatives of olig ouridylates pU(n) (n = 3, 4, 6, 12) bearing 5'-P-32-label was studied. Complexes of these derivatives with 80S ribosomes where codon-anticod on interaction took place either in P-site (in the case of pU3-and pU4 -derivatives), or in P- and A-site simultaneously (in the case of pU6- and pU12-derivatives) were obtained in the presence of Phe-tRNA(Phe). All the reagents modified only the 40S subunit. The extent of 18S rRN A modification by pU3-, pU4-, pU6- and pU12-derivatives as a fraction of the total modification extent of 18S rRNA and proteins in the 40S s ubunit equaled 96, 93, 24 and 4%, respectively. The pU4-derivative was covalently attached at positions 976-1061 and 1058-1164 and pU12-deri vative was covalently attached within regions 976-1061, 1058-1164, 593 -673 and 1748-1869 of the 18S rRNA. By means of the primer extension t echnique, modified bases in 18S rRNA were determined to be: A-1023, C- 1026, A-1027, A-1058, G-1059 for pU3- and pU4-derivatives and A-1058 f or pU6-derivative.