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 intimately with the signal-recognition particle RNA (SRP RNA) and serves as an important structural and functional component of the SRP. We investigated the structure and RNA-binding activity of the human SRP19 protein by the use of comparative sequence analysis, high-stringency structure prediction, proteolytic susceptibility, and site-directed mutagenesis. SRP19 was found to consist of two distinct regions (called N-terminal and C-terminal regions) that are separated by a boundary of approximately 12-15 amino acid residues. Both regions contain an alpha-helix and several beta-strands that are connected by loops or turns. In agreement with the hypothetical model, proteolytic susceptibility demonstrated the predominant accessibility of two sites: one in a surface loop of the N-terminal region (YLNNKKTIAEGR33), and another site in the C-terminal tail at residues L129 and E133. The RNA-binding activities of mutant polypeptides with changes of conserved lysines and arginines (mutants K27Q, R33Q and R34Q) demonstrated that the proteolytically accessible loop of the N-terminal region is in direct contact with the SRP RNA. In contrast, alteration of a certain basic amino acid residues in 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 regions, had no effect on the RNA-binding ability. The structural model that emerges from our data is thematically similar to that of ribosomal protein S5, the N-domain of which contains a loop motif believed to interact with double-stranded RNA. The presence of a similar structural feature in protein SRP19 has significant implications for the structure and function of the SRP19-RNA complex.
Black
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Department of Molecular Biology, The University of Texas Health Science Center at Tyler, 75710, USA.
Gowda
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Chittenden
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Walker
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KP
Zwieb
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D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010944:Plants; D011485:Protein Binding; D017433:Protein Structure, Secondary; D012313:RNA / Q000378:metabolism*; D012441:Saccharomyces cerevisiae; D017421:Sequence Analysis; D018271:Signal Recognition Particle / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D018271:Signal Recognition Particle; D012313:RNA