Fig. 1.

Sequence analysis of peptide-RNA (oligo)nucleotide cross-links derived from human spliceosomal complex U4 specific 15.5K protein cross-linked to U4 snRNA. A, MS/MS spectrum of m/z 874.3856²⁺ annotated as LLDLVQQSCNYK (positions 22–33) cross-linked to nucleotide U. The peptide fragment signals shifted by (U-H₃PO₄) or (226 Da) are indicated by #. B, MS/MS spectrum of m/z 663.9234³⁺ annotated as LLDLVQQSCNYK (positions 22–33) cross-linked to nucleotide U with the additional mass of 152 Da. The peptide fragment signals shifted by [(U-H₃PO₄)+152 Da] or (378 Da) are indicated by #. C, MS/MS spectrum of m/z 954.8976²⁺ annotated as LLDLVQQSCNYK (positions 22–33) cross-linked to nucleotide ¹³U with the additional mass of 152 Da. Fragment ions shifted by an additional mass of 387.0858 Da are indicated by #. Signals corresponding to uracil fragments with the 152 Da increment are annotated as U', (y_r2-y_r1) and (y_r2-w_r1). All signals corresponding to RNA and peptide-RNA adducts are highlighted by bold letter. RNA oligonucleotide fragmentation nomenclature is according to McLuckey et al. (61). A proposed schematic illustration of the peptide-RNA cross-link along with cross-link product of +152 Da is shown on the right. Cysteine as the cross-linked amino acid identified by shift in the y ion series (starting from the y₄ ion), is highlighted by bold letter in the peptide sequence. y-type fragmentation of RNA is referred to as U', w_r1, y_r1, y_r2. Positions of ¹³C on UTP are marked by orange circles in lower panel.