Click chemistry generated model DNA-peptide heteroconjugates as tools for mass spectrometry.
UV cross-linking of nucleic acids to proteins in combination with mass spectrometry is a powerful technique to identify proteins, peptides, and the amino acids involved in intermolecular interactions within nucleic acid-protein complexes. However, the mass spectrometric identification of cross-linked nucleic acid-protein heteroconjugates in complex mixtures and MS/MS characterization of the specific sites of cross-linking is extremely challenging. As a tool for the optimization of sample preparation, ionization, fragmentation, and detection by mass spectrometry, novel synthetic DNA-peptide heteroconjugates were generated to act as mimics of UV cross-linked heteroconjugates. Click chemistry was employed to cross-link peptides to DNA oligonucleotides. These heteroconjugates were fully characterized by high resolution FTICR mass spectrometry and by collision-induced dissociation (CID) following nuclease P1 digestion of the DNA moiety to a single nucleotide monophosphate. This allowed the exact site of the cross-linking within the peptide to be unambiguously assigned. These synthetic DNA-peptide heteroconjugates have the potential to be of use for a variety of applications that involve DNA-peptide heteroconjugates.
Institute of Cell Biology, School of Biological Sciences, University of Edinburgh , Edinburgh EH9 3FF, Scotland, United Kingdom.
Fiona J
FJ
Flett
School of Chemistry, University of Edinburgh , Edinburgh EH9 3FJ, Scotland, United Kingdom.
Jeffrey G A
JG
Walton
School of Chemistry, University of Edinburgh , Edinburgh EH9 3FJ, Scotland, United Kingdom.
C Logan
CL
Mackay
Institute of Cell Biology, School of Biological Sciences, University of Edinburgh , Edinburgh EH9 3FF, Scotland, United Kingdom.
Heidrun
H
Interthal
D002384:Catalysis; D057930:Click Chemistry; D003300:Copper / Q000737:chemistry; D004247:DNA / Q000737:chemistry*; D013058:Mass Spectrometry; D015394:Molecular Structure; D010455:Peptides / Q000737:chemistry*
D010455:Peptides; D003300:Copper; D004247:DNA