Human ribosomal protein S13 is a structural element of the small ribosome subunit. It is homologous to the eubacterial ribosomal protein S15 and additionally possesses an extended N-terminal region, characteristic of the S15p family in eukaryotes and archaea. In the present study, we investigated the binding of the recombinant ribosomal protein S13 and its mutant forms containing deletions or amino acid substitutions in different regions to an RNA transcript representing a fragment of the central domain of the 18S rRNA. It was shown that substitution of ultra-conservative residues H101 and D108, as well as deletions of either 29 C-terminal or 27 N-terminal residues, substantially reduced the protein affinity to the RNA transcript. Deletion of 54 C-terminal or 80 N-terminal residues rendered the protein fully incapable of RNA binding. Using a footprinting assay, we identified those sites in the RNA transcript that change their accessibility to hydroxyl radicals as a result of binding either full-length protein S13 or its mutant lacking 27 N-terminal residues. It was shown that these sites are located mainly in the H22 helix of the 18S rRNA and near its junction with the H20 helix and that they largely correspond to rRNA contacts with the conserved part of the protein. Thus, the binding of the ribosomal protein S13 to the 18S rRNA is provided mainly by conserved motifs of the protein corresponding to those motifs in its eubacterial homologue that are involved in the interaction with the 16S rRNA in the 30S subunit. The role of the N-terminal region of S13 in its binding to the central domain of the 18S rRNA is discussed.