Analysis of Protein-protein Interaction Interface between Yeast Mitochondrial Proteins Rim1 and Pif1 Using Chemical Cross-linking Mass Spectrometry.

ABSTRACT: Defining protein-protein contacts is a challenging problem and cross-linking is a promising solution. Here, we present a case of mitochondrial single strand binding protein Rim1 and helicase Pif1, an interaction first observed in immuno-affinity pull-down from yeast cells using Pif1 bait. We found that only the short succinimidyl-diazirine cross-linker or formaldehyde captured the interaction between recombinant Rim1 and Pif1. In addition, Pif1 needed to be stripped of its N-terminal and C-terminal domains, and Rim1's C-terminus needed to be modified for the cross-linked product to become visible. Our report is an example of a non-trivial analysis, where a previously identified stable interaction escapes initial capture with cross-linking agents and requires substantial modification to rec

SUBMITTER: Zybailov B 

PROVIDER: S-EPMC4722957 | biostudies-literature | 2015 Nov

REPOSITORIES: biostudies-literature