Interaction of ribosomal proteins S6, S8, S15 and S18 with the central domain of 16 S ribosomal RNA from Escherichia coli.
The co-operative interaction of 30 S ribosomal subunit proteins S6, S8, S15 and S18 with 16 S ribosomal RNA from Escherichia coli was studied by (1) determining how the binding of each protein is influenced by the others and (2) characterizing a series of protein-rRNA fragment complexes. Whereas S8 and S15 are known to associate independently with the 16 S rRNA, binding of S18 depended upon S8 and S15, and binding of S6 was found to require S8, S15 and S18. Ribonucleoprotein (RNP) fragments were derived from the S8-, S8/S15- and S6/S8/S15/S18-16 S rRNA complexes by partial RNase hydrolysis and isolated by electrophoresis through Mg2+-containing polyacrylamide gels or by centrifugation through sucrose gradients. Identification of the proteins associated with each RNP by gel electrophoresis in the presence of sodium dodecyl sulfate demonstrated the presence of S8, S8 + S15 and S6 + S8 + S15 + S18 in the corresponding fragment complexes. Analysis of the rRNA components of the RNP particles confirmed that S8 was bound to nucleotides 583 to 605 and 624 to 653, and that S8 and S15 were associated with nucleotides 583 to 605, 624 to 672 and 733 to 757. Proteins S6, S8, S15 and S18 were shown to protect nucleotides 563 to 605, 624 to 680, 702 to 770, 818 to 839 and 844 to 891, which span the entire central domain of the 16 S rRNA molecule (nucleotides 560 to 890). The binding site for each protein contains helical elements as well as single-stranded internal loops ranging in size from a single bulged nucleotide to 20 bases. Three terminal loops and one stem-loop structure within the central domain of the 16 S rRNA were not protected in the four-protein complex. Interestingly, bases within or very close to these unprotected regions have been shown to be accessible to chemical and enzymatic probes in 30 S subunits but not in 70 S ribosomes. Furthermore, nucleotides adjacent to one of the unprotected loops have been cross-linked to a region near the 3' end of 16 S rRNA. Our observations and those of others suggest that the bases in this domain that are not sequestered by interactions with S6, S8, S15 or S18 play a role involved in subunit association or in tertiary interactions between portions of the rRNA chain that are distant from one-another in the primary structure.(ABSTRACT TRUNCATED AT 400 WORDS)
D001483:Base Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000032:analysis*; D046911:Macromolecular Substances; D009690:Nucleic Acid Conformation; D012329:RNA, Bacterial / Q000378:metabolism*; D012335:RNA, Ribosomal / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000032:analysis; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins / Q000378:metabolism*
D046911:Macromolecular Substances; D012329:RNA, Bacterial; D012335:RNA, Ribosomal; D012261:Ribonucleoproteins; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins; C036004:ribosomal protein S15; C041184:ribosomal protein S18; C030254:ribosomal protein S8