Crosslinking of an iodo-uridine-RNA hairpin to a single site on the human U1A N-terminal RNA binding domain.
The N-terminal RNA binding domain (RBD) of the human U1A snRNP protein binds tightly and specifically to an RNA hairpin that contains a 10-nucleotide loop. The protein is one of a class of RNA binding proteins that adopts a beta alpha beta beta alpha beta global fold, which in turn forms a four-stranded antiparallel beta-sheet. This sheet forms the primary binding surface for the RNA, as shown by the crosslinking results described here, and in more detail by a recently described co-crystal of this RBD with an RNA hairpin (Oubridge C, et al., 1994, Nature 372:432-438). The RNA hairpin sequence used in the crosslinking experiments, containing 5-iodo-uridine, is a variant of the normal U1 snRNA sequence which is able to form a crosslink with the protein, in contrast to the wild-type sequence, which does not. This single uridine substitution in the 10-nucleotide loop is the site of cross-linking to one tyrosine (Tyr 13) in the beta 1 strand of the U1A N-terminal RBD. This same uridine is also crosslinked to a mutant Tyr 13 Phe RBD, at this Phe 13 substitution.
D001483:Base Sequence; D003432:Cross-Linking Reagents; D006801:Humans; D007065:Idoxuridine / Q000031:analogs & derivatives; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D010777:Photochemistry; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D012342:RNA, Small Nuclear / Q000737:chemistry / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017412:Ribonucleoprotein, U1 Small Nuclear / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017421:Sequence Analysis; D013329:Structure-Activity Relationship
D003432:Cross-Linking Reagents; D012342:RNA, Small Nuclear; D016601:RNA-Binding Proteins; D017412:Ribonucleoprotein, U1 Small Nuclear; C075339:U1A protein; C035991:5-iodouridine; D007065:Idoxuridine