A photocleavable and mass spectrometry identifiable cross-linker for protein interaction studies.
In this paper, we present the results of proof-of-concept experiments using a novel photocleavable and mass spectrometry identifiable cross-linker pcPIR (photocleavable protein interaction reporter). pcPIR can be dissociated under UV irradiation either off- or online before the introduction to the mass spectrometers. Photo dissociation of cross-linkers is different from either the gas phase or the chemical cleavage of cross-linkers. Different types of cross-links can be identified using the pcPIR mass relationships, where the mass of cross-linked precursor equals the sum of the masses of the released products and reporter. Since pcPIR is cleaved prior to the entrance to the mass spectrometer, the released peptides are available to be sequenced with routine collision-induced dissociation (CID) MS/MS experiments and database search algorithms. In this report, the pcPIR strategy of identifying the cross-linked peptides with on- and off-line photocleavage coupled with novel targeted data dependent LC-MS/MS is demonstrated with the use of standard peptides, bovine serum albumin (BSA), and human hemoglobin tetramer protein complex.
Yang
Li
L
Department of Chemistry, Washington State University, Pullman, Washington 99164, USA.
Tang
Xiaoting
X
Weisbrod
Chad R
CR
Munske
Gerhard R
GR
Eng
Jimmy K
JK
von Haller
Priska D
PD
Kaiser
Nathan K
NK
Bruce
James E
JE
D000818:Animals; D002417:Cattle; D003432:Cross-Linking Reagents / Q000138:chemical synthesis* / Q000737:chemistry; D006801:Humans; D008027:Light; D013058:Mass Spectrometry; D015394:Molecular Structure; D010777:Photochemistry; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry*
D003432:Cross-Linking Reagents; D011506:Proteins