TY - JOUR
T1 - Hidden information on protein function in censuses of proteome foldedness
AU - Cox, Dezerae
AU - Ang, Ching Seng
AU - Nillegoda, Nadinath B.
AU - Reid, Gavin E.
AU - Hatters, Danny M.
N1 - Funding Information:
We thank Professor Pierre Goloubinoff from University of Lausanne for provision of unfolding data of DNAJB1 (in Supplementary Fig.?6C). We thank Professors Paul Gooley and Heath Ecroyd for helpful discussions and careful reading of the manuscript. We also thank Dr. Yuning Hong (La Trobe University) for providing TPE-MI, and the Bio21 Melbourne Mass Spectrometry and Proteomics facility. This work was funded by grants National Health and Medical Research Council APP1161803 (D.M.H.) and Australian Research Council DP170103093 (D.M.H. and G.E.R.).
Funding Information:
We thank Professor Pierre Goloubinoff from University of Lausanne for provision of unfolding data of DNAJB1 (in Supplementary Fig. ). We thank Professors Paul Gooley and Heath Ecroyd for helpful discussions and careful reading of the manuscript. We also thank Dr. Yuning Hong (La Trobe University) for providing TPE-MI, and the Bio21 Melbourne Mass Spectrometry and Proteomics facility. This work was funded by grants National Health and Medical Research Council APP1161803 (D.M.H.) and Australian Research Council DP170103093 (D.M.H. and G.E.R.).
Publisher Copyright:
© 2022, The Author(s).
PY - 2022/12
Y1 - 2022/12
N2 - Methods that assay protein foldedness with proteomics have generated censuses of apparent protein folding stabilities in biological milieu. However, different censuses poorly correlate with each other. Here, we show that the reason for this is that methods targeting foldedness through monitoring amino acid sidechain reactivity also detect changes in conformation and ligand binding, which can be a substantial fraction of the data. We show that the reactivity of only one quarter of cysteine or methionine sidechains in proteins in a urea denaturation curve of mammalian cell lysate can be confidently explained by a two-state unfolding isotherm. Contrary to that expected from unfolding, up to one third of the cysteines decreased reactivity. These cysteines were enriched in proteins with functions relating to unfolded protein stress. One protein, chaperone HSPA8, displayed changes arising from ligand and cofactor binding. Unmasking this hidden information using the approaches outlined here should improve efforts to understand both folding and the remodeling of protein function directly in complex biological settings.
AB - Methods that assay protein foldedness with proteomics have generated censuses of apparent protein folding stabilities in biological milieu. However, different censuses poorly correlate with each other. Here, we show that the reason for this is that methods targeting foldedness through monitoring amino acid sidechain reactivity also detect changes in conformation and ligand binding, which can be a substantial fraction of the data. We show that the reactivity of only one quarter of cysteine or methionine sidechains in proteins in a urea denaturation curve of mammalian cell lysate can be confidently explained by a two-state unfolding isotherm. Contrary to that expected from unfolding, up to one third of the cysteines decreased reactivity. These cysteines were enriched in proteins with functions relating to unfolded protein stress. One protein, chaperone HSPA8, displayed changes arising from ligand and cofactor binding. Unmasking this hidden information using the approaches outlined here should improve efforts to understand both folding and the remodeling of protein function directly in complex biological settings.
UR - https://www.scopus.com/pages/publications/85128260889
U2 - 10.1038/s41467-022-29661-2
DO - 10.1038/s41467-022-29661-2
M3 - Article
C2 - 35422070
AN - SCOPUS:85128260889
SN - 2041-1723
VL - 13
JO - Nature Communications
JF - Nature Communications
IS - 1
M1 - 1992
ER -