Open search strategy for inferring the masses of cross-link adducts

ABSTRACT: Development of new reagents for protein cross-linking is continually ongoing. The chemical formulas of the linker adducts formed by these reagents are usually deduced from expert knowledge, and then validated by mass spectrometry. Clearly, it would have been more rigorous if the adduct masses are inferred directly from the data without any prior assumption on the chemistries. Unfortunately, the analysis tools that are currently available to detect chemical modifications on linear peptides are not applicable to the case of two cross-linked peptides. Here we show that an adaptation of the open search strategy that works on linear peptides can be used to study cross-link modifications between pairs of peptides. We benchmark our approach by correctly inferring the linker masses of two well-known reagents - DSS and formaldehyde - to accuracies of a few parts per million. We then investigate the cross-linking chemistries of two poorly characterized reagents - EMCS and glutaraldehyde. In the case of EMCS, we find that the expected cross-linking chemistry is accompanied by a competing chemistry that targets other amino acid types. In the case of glutaraldehyde, we find that the chemical formula of the dominant linker is C5H4, which indicates a ringed aromatic structure. These results demonstrate that, at very little effort, our approach can yield non-trivial insights to facilitate the studies of new cross-linkers.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Bos Taurus (bovine)

SUBMITTER: Moriya Slavin  

LAB HEAD: Nir Kalisman

PROVIDER: PXD020704 | Pride | 2021-09-09

REPOSITORIES: Pride