title: Mixed-isotope labeling with LC-IMS-MS for characterization of protein-protein interactions by chemical cross-linking.
abstract: Chemical cross-linking of proteins followed by proteolysis and mass spectrometric analysis of the resulting cross-linked peptides provides powerful insight into the quaternary structure of protein complexes. Mixed-isotope cross-linking (a method for distinguishing intermolecular cross-links) was coupled with liquid chromatography, ion mobility spectrometry and mass spectrometry (LC-IMS-MS) to provide an additional separation dimension to the traditional cross-linking approach. This method produced multiplet m/z peaks that are aligned in the IMS drift time dimension and serve as signatures of intermolecular cross-linked peptides. We developed an informatics tool to use the amino acid sequence information inherent in the multiplet spacing for accurate identification of the cross-linked peptides. Because of the separation of cross-linked and non-cross-linked peptides in drift time, our LC-IMS-MS approach was able to confidently detect more intermolecular cross-linked peptides than LC-MS alone.
authors: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA 99352, USA.
authors: Eric D
authors: ED
authors: Merkley
authors: Erin S
authors: ES
authors: Baker
authors: Kevin L
authors: KL
authors: Crowell
authors: Daniel J
authors: DJ
authors: Orton
authors: Thomas
authors: T
authors: Taverner
authors: Charles
authors: C
authors: Ansong
authors: Yehia M
authors: YM
authors: Ibrahim
authors: Meagan C
authors: MC
authors: Burnet
authors: John R
authors: JR
authors: Cort
authors: Gordon A
authors: GA
authors: Anderson
authors: Richard D
authors: RD
authors: Smith
authors: Joshua N
authors: JN
authors: Adkins
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D002853:Chromatography, Liquid / Q000379:methods; D003432:Cross-Linking Reagents / Q000737:chemistry*; D007553:Isotope Labeling / Q000379:methods; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis* / Q000378:metabolism; D011487:Protein Conformation; D025941:Protein Interaction Mapping / Q000379:methods*; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020539:Sequence Analysis, Protein / Q000379:methods; D020592:Shewanella / Q000737:chemistry / Q000378:metabolism; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D010455:Peptides; D011506:Proteins; D011994:Recombinant Proteins