Targeted cross-linking-mass spectrometry determines vicinal interactomes within heterogeneous RNP complexes.

Abstract:

:Proteomic and RNomic approaches have identified many components of different ribonucleoprotein particles (RNPs), yet still little is known about the organization and protein proximities within these heterogeneous and highly dynamic complexes. Here we describe a targeted cross-linking approach, which combines cross-linking from a known anchor site with affinity purification and mass spectrometry (MS) to identify the changing vicinity interactomes along RNP maturation pathways. Our method confines the reaction radius of a heterobifunctional cross-linker to a specific interaction surface, increasing the probability to capture low abundance conformations and transient vicinal interactors too infrequent for identification by traditional cross-linking-MS approaches, and determine protein proximities within RNPs. Applying the method to two conserved RNA-associated complexes in Saccharomyces cerevisae, the mRNA export receptor Mex67:Mtr2 and the pre-ribosomal Nop7 subcomplex, we identified dynamic vicinal interactomes within those complexes and along their changing pathway milieu. Our results therefore show that this method provides a new tool to study the changing spatial organization of heterogeneous dynamic RNP complexes.

journal_name

Nucleic Acids Res

journal_title

Nucleic acids research

authors

Trahan C,Oeffinger M

doi

10.1093/nar/gkv1366

subject

Has Abstract

pub_date

2016-02-18 00:00:00

pages

1354-69

issue

3

eissn

0305-1048

issn

1362-4962

pii

gkv1366

journal_volume

44

pub_type

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