UV (lambda = 254 nm) irradiation of bacteriophage MS2 or its treatment with bisulfite induce covalent crosslinkage of the RNA to the coat protein. epilsonN-(2-oxopyrimidyl-4)-lysine was found in the phage hydrolysates after either type of treatment. An equimolar mixture of 0-methylhydroxylamine and bisulfite causes complete disappearance of the cross-links. This led to the conclusion that one of the factors responsible for the UV-induced polynucleotide-protein crosslinkage and the main factor in treatment with bisulfite is substitution of the exocyclic amino group of the activated cytosine nucleus by the lysine residue epilson-amino group of the protein.

Ultraviolet irradiation (lambda = 254 nm) of ternary complexes of E. coli 70 S ribosomes with poly(U) and either Phe-tRNAPhe (in the A-site) or NAcPhe-tRNAPhe (in the P-site) effectively induces covalent linking of tRNA with a limited number of ribosomal proteins. The data obtained indicate that in both sites tRNA is in contact with proteins of both 30 S and 50 S subunits (S5, S7, S9, S10, L2, L6 and L16 proteins in the A-site and S7, S9, S11, L2, L4, L7/L12 and L27 proteins in the P-site). Similar sets of proteins are in contact with total aminoacyl-tRNA and N-acetylaminoacyl-tRNA. However, here no contacts of tRNA in the P-site with the S7 and L25/S17 proteins were revealed, whereas in the A-site total aminoacyl-tRNA contacts L7/L12. Proteins S9, L2 and, probably, S7 and L7/L12 are common to both sites.

Direct contacts between 16-S RNA and split proteins S2, S3, S 5 , S14 and S21 inside the 30-S subunit of Eschevichia coli ribosomes were evidenced by the formation of ultraviolet-induced (A = 254 nni) RNA-protein cross-links. 30-S subunits were reassembled from core particles and a mixture of split proteins containing in each case a single '251-labelled protein. All the proteins tested are cross-linked as a result of a single-hit process; proteins S3 and S21 were cross-linked at the highest rate.The essential step in the study of nucleoproteins is detection of interactions between components of the complex.The evidence for direct RNA-protein contacts in ribosomes of Eschevichia coli was presented for a number of proteins [l -31. They belong mostly to a group of proteins which do not readily dissociate from 16-S RNA upon incubation of 30-S subparticles at high concentrations of LiCl or CsCl and which form specific RNA . protein complexes (core proteins) [4,5]. At the same time, there was no evidence for the existence of direct RNA-protein contacts in the 30-S subparticle for a number of so-called split proteins that readily dissociate from the ribosome at high concentrations of LiCl or CsCl and do not form strong complexes with 16-S RNA.Only recently [6] Hochkeppel and Craven showed that some split proteins are able to bind independently to 16-S RNA prepared by acetic acid urea extraction. They presented some evidence that six of these proteins (S3, S5, S9, S11, S12 and SlS) bind site-specifically to such isolated 16-S RNA. But the stability and specificity of nucleoproteins are conditioned by cooperative interactions. Therefore the data, obtained for incomplete complexes, seem not to be very conclusive.For detection of contacts between a number of split proteins (S2, S3, S5, S14 and S21) and 16-S RNA in the complete 30-S subunit of E. coli ribosomes we have used ultraviolet-induced cross-links. This method is adequate for fixation of polynucleotideprotein contacts in intact nucleoproteins [7,8] including ribosomes [9-121. 30-S subunits were reconstituted from a mixture of core subparticles and split proteins, containing in each case a single "'Ilabelled protein component. The reconstituted subunits were irradiated with ultraviolet light (i = 253.7 nm, 2.5-25 x lo5 quanta per subparticle) and the formation of cross-links was monitored by analysis of cosedimentation of 16-S RNA and the labelled protein in the presence of sodium dodecyl sulphate and EDTA.It was shown that proteins S3 and S21 cross-linked well with 16-S RNA, whereas proteins S2, S5 and S14 cross-linked under the same conditions to a much lesser extent. MATERIALS AND METHODS30-S subunits of E. coli MRE 600 ribosomes were prepared by zonal centrifugation [I 31. Individual ribosomal proteins were isolated according to [14]. 100-300 pg of each isolated protein were labelled with 1-3 pCi of Na'251 (Amersham, 18 mCi/pg) by the method of Ball et al. [15] with a slight modification [16]. The purity of the labelled proteins was checked by gel electrophoresis...

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