Chemical cross-linking and mass spectrometry to determine the subunit interaction network in a recombinant human SAGA HAT subcomplex

Protein Science : a Publication of the Protein Society
Nha-Thi Nguyen-HuynhEmmanuelle Leize-Wagner

Abstract

Understanding the way how proteins interact with each other to form transient or stable protein complexes is a key aspect in structural biology. In this study, we combined chemical cross-linking with mass spectrometry to determine the binding stoichiometry and map the protein-protein interaction network of a human SAGA HAT subcomplex. MALDI-MS equipped with high mass detection was used to follow the cross-linking reaction using bis[sulfosuccinimidyl] suberate (BS3) and confirm the heterotetrameric stoichiometry of the specific stabilized subcomplex. Cross-linking with isotopically labeled BS3 d0-d4 followed by trypsin digestion allowed the identification of intra- and intercross-linked peptides using two dedicated search engines: pLink and xQuest. The identified interlinked peptides suggest a strong network of interaction between GCN5, ADA2B and ADA3 subunits; SGF29 is interacting with GCN5 and ADA3 but not with ADA2B. These restraint data were combined to molecular modeling and a low-resolution interacting model for the human SAGA HAT subcomplex could be proposed, illustrating the potential of an integrative strategy using cross-linking and mass spectrometry for addressing the structural architecture of multiprotein complexes.

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Jul 28, 2016·Analytical Chemistry·Sven H GieseJuri Rappsilber
Jun 30, 2018·Mass Spectrometry Reviews·Kevin TitecaSven Eyckerman
Jul 29, 2018·The Journal of Biological Chemistry·Chitvan MittalMichael A Shogren-Knaak
Oct 3, 2020·Genes & Development·Jelly H M Soffers, Jerry L Workman
Sep 6, 2020·Biochimica Et Biophysica Acta. Gene Regulatory Mechanisms·Jose M Espinola-Lopez, Song Tan
Aug 3, 2020·Biochimica Et Biophysica Acta. Gene Regulatory Mechanisms·Dominique HelmlingerLászló Tora
Nov 1, 2020·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Marco Giampà, Elvira Sgobba
Aug 12, 2017·Journal of the American Society for Mass Spectrometry·Daniel S ZiemianowiczDavid C Schriemer

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