Analysis of protein--RNA interactions within Ro ribonucleoprotein complexes.
The interactions between Ro and La proteins and hY RNAs have been analysed. The binding site for the 60 kDa Ro protein on hY RNAs is shown to be the terminal part of the base paired stem structure, which contains the most highly conserved sequence among hY RNAs. The bulged C-residue within this region plays an important role in the recognition by this protein. The same regions of hY RNAs are essential for the association of the 52 kDa Ro protein with the RNAs, strongly suggesting that the 60 kDa Ro protein is required for the 52 kDa Ro protein to bind, presumably via protein-protein interactions, to Ro RNPs. The binding site for the La protein on hY RNAs is shown to be the oligouridylate stretch near the 3'-end of the RNAs, which is also recognized when additional nucleotides flank this motif at the 3'-side. Additional sequence elements in hY3 and hY5, but not in hY1, are bound by the La protein as well. Deletion mutagenesis showed that the RNP motif, previously identified in many ribonucleoprotein (RNP) proteins and in some cases shown to be almost sufficient for the interaction with RNA, of both the 60 kDa Ro and the La protein are not sufficient for the interaction with hY RNAs. Substantial parts of these proteins flanking the RNP motif are needed as well. It is likely that they stabilize the correct conformation of the RNP motif for RNA binding.
Pruijn
G J
GJ
Department of Biochemistry, University of Nijmegen, The Netherlands.
Slobbe
R L
RL
van Venrooij
W J
WJ
D001482:Base Composition / Q000502:physiology; D001483:Base Sequence; D001665:Binding Sites / Q000502:physiology; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D006367:HeLa Cells; D006801:Humans; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011072:Poly U / Q000378:metabolism; D012342:RNA, Small Nuclear / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012261:Ribonucleoproteins / Q000378:metabolism*; D017411:Ribonucleoproteins, Small Nuclear
D046911:Macromolecular Substances; D012342:RNA, Small Nuclear; D011994:Recombinant Proteins; D012261:Ribonucleoproteins; D017411:Ribonucleoproteins, Small Nuclear; D011072:Poly U