IDENTIFICATION OF AN RNA-BINDING-LOOP IN THE N-TERMINAL REGION OF SIGNAL-RECOGNITION-PARTICLE PROTEIN SRP19

Protein SRP19 is a 144-amino-acid polypeptide that associates intimate ly with the signal-recognition particle RNA (SRP RNA) and serves as an important structural and functional component of the SRP. We investig ated the structure and RNA-binding activity of the human SRP19 protein by the use of comparative sequence analysis, high-stringency structur e prediction, proteolytic susceptibility, and site-directed mutagenesi s. SRP19 was found to consist of two distinct regions (called N-termin al and C-terminal regions) that are separated by a boundary of approxi mately 12-15 amino acid residues. Both regions contain an alpha-helix and several beta-strands that are connected by loops or turns In agree ment with the hypothetical model, proteolytic susceptibility demonstra ted the predominant accessibility of two sites: one in a surface loop of the N-terminal region (YLNNKKTIAEGR33), and another site in the C-t erminal tail at residues L129 and E133. The RNA-binding activities of mutant polypeptides with changes of conserved lysines and arginines (m utants K27Q, R33Q and R34Q) demonstrated that the proteolytically acce ssible loop of the N-terminal region is in direct contact with the SRP RNA. Ln contrast, alteration of a certain basic amino acid residues i n the C-terminal region (R83, K116 and R118), as well as a deletion of four amino acid residues located at the boundary between the two regi ons, had no effect on the RNA-binding ability. The structural model th at emerges from our data is thematically similar to that of ribosomal protein S5, the N-domain of which contains a loop motif believed to in teract with double-stranded RNA. The presence of a similar structural feature in protein SRP19 has significant implications for the structur e and function of the SRP19-RNA complex.