"Zero-length" cross-linking in solid state as an approach for analysis of protein-protein interactions.

ABSTRACT: We have developed a new approach for the analysis of interacting interfaces in protein complexes and protein quaternary structure based on cross-linking in the solid state. Protein complexes are freeze-dried under vacuum, and cross-links are introduced in the solid phase by dehydrating the protein in a nonaqueous solvent creating peptide bonds between amino and carboxyl groups of the interacting peptides. Cross-linked proteins are digested into peptides with trypsin in both H2(16)O and H(2)18O and then readily distinguished in mass spectra by characteristic 8 atomic mass unit (amu) shifts reflecting incorporation of two 18O atoms into each C terminus of proteolytic peptides. Computer analysis of mass spectrometry (MS) and MS/MS data is used to identify the cross-linked peptides. We demonst

SUBMITTER: El-Shafey A 

PROVIDER: S-EPMC2249764 | biostudies-literature | 2006 Mar

REPOSITORIES: biostudies-literature