MESSENGER-RNA RECOGNITION BY FRAGMENTS OF RIBOSOMAL-PROTEIN S4

Ribosomal protein S4 from Escherichia coli binds a large domain of 16 S ribosomal RNA and also a pseudoknot structure in the alpha operon mR NA, where it represses its own synthesis. No similarity between the tw o RNA binding sites has been detected. To find out whether separate pr otein regions are responsible for rRNA and mRNA recognition, proteins with N-terminal or C-terminal deletions have been overexpressed and pu rified. Protein-mRNA interactions were detected by (i) a nitrocellulos e filter binding assay, (ii) inhibition of primer extension by reverse transcriptase, and (iii) a gel shift assay, Circular dichroism spectr a were taken to determine whether the proteins adopted stable secondar y structures, From these studies it is concluded that amino acids 48-1 04 make specific contacts with the mRNA, although residues 105-177 (ou t of 205) are required to observe the same toeprint pattern as full-le ngth protein and may stabilize a specific portion of the mRNA structur e, These results parallel ribosomal RNA binding properties of similar fragments (Conrad, R. C. and Craven, G. R. (1987) Nucleic Acids Res. 1 5, 10331-10343, and references therein). It appears that the same prot ein domain is responsible for both mRNA and rRNA binding activities.