Photoinduced crosslinkage, in situ, of Escherichia coli 30S ribosomal proteins to 16S rRNA: identification of crosslinked proteins and relations between reactivity and ribosome structure

Abstract

The kinetics of photoinduced cross-linkage of E. coli 30S ribosomal proteins to the 16S rRNA molecule in the intact E. coli 30S ribosomal subunit was studied. All of the 30S ribosomal proteins become cross-linked to the 16S rRNA before changes in the sedimentation characteristics of the 30S ribosomal subunit can be detected. The proteins exhibit different reactivities in the cross-linkage reaction. One group of proteins, S3, S7-S9, S11, S12 and S15-S19, is cross-linked to the 16S rRNA by single-hit kinetics or by photoprocesses of nonunity but low multiplicities. A 2nd group of proteins, S1, S2, S4-S6, S10, S13, S14 and S21, is cross-linked to the 16S rRNA by photoprocesses of a complex nature. A comparison of these data with other properties of the individual 30S ribosomal proteins related to ribosome structure indicated that most of the 30S ribosomal proteins cross-linked to the 16S rRNA by photoprocesses of low multiplicities were classified rRNA-binding proteins by nonphotochemical methods and most of the proteins cross-linked to the 16S rRNA by photoprocesses of large multiplicities were classified as non-binding proteins. There were certain exceptions to these correlations. Proteins S4 and S20, both RNA-binding proteins, become cross-linked to the 16S rRNA by photoprocesses of large multiplicities and proteins S3, S11, S12 and S18, none of which were classified RNA-binding proteins, exhibited low multiplicities in the cross-linkage reaction. All of these exceptions could be explained in terms of limitations inherent in the photochemical methods used in this study and in other types of methods that were used to study RNA-protein interactions in the 30S ribosomal subunit. Labile RNA-protein cross-links are apparently present in the UV-irradiated 30S ribosomal subunits and neither peptide-bond cleavage nor photoinduced modification of the charged side-chain groups in the ribosomal proteins apparently accompanied the cross-linkage reaction. Some photoinduced RNA-chain breakage might have occurred.