TY - JOUR
T1 - Antibody for Serine 65 Phosphorylated Ubiquitin Identifies PLK1-Mediated Phosphorylation of Mitotic Proteins and APC1
AU - Mann, Guy
AU - Sulkshane, Prasad
AU - Sadhu, Pradeep
AU - Ziv, Tamar
AU - Glickman, Michael H.
AU - Brik, Ashraf
N1 - Funding Information:
This work was partially funded by grant 755/19 from the Israel Science Foundation (M.H.G.), 76251-12-9/19 (ZN3457) from the Ministry of Science and Culture of Lower Saxony for research on mitochondria (M.H.G.) and grants 831783 from the European Research Council (ERC) under the European Union’s Horizon-2020 research and innovation program (grant agreement no. [831783]) (A.B.). This work is a product of COST action CA20113 ProteoCure. A.B. is the Jordan and Irene Tark Academic Chair, M.H.G. is the Israel Isaac and Natalia Kudish Academic Chair.
Publisher Copyright:
© 2022 by the authors.
PY - 2022/8/1
Y1 - 2022/8/1
N2 - Deciphering the protein posttranslational modification (PTM) code is one of the greatest biochemical challenges of our time. Phosphorylation and ubiquitylation are key PTMs that dictate protein function, recognition, sub-cellular localization, stability, turnover and fate. Hence, failures in their regulation leads to various disease. Chemical protein synthesis allows preparation of ubiquitinated and phosphorylated proteins to study their biochemical properties in great detail. However, monitoring these modifications in intact cells or in cell extracts mostly depends on antibodies, which often have off-target binding. Here, we report that the most widely used antibody for ubiquitin (Ub) phosphorylated at serine 65 (pUb) has significant off-targets that appear during mitosis. These off-targets are connected to polo-like kinase 1 (PLK1) mediated phosphorylation of cell cycle-related proteins and the anaphase promoting complex subunit 1 (APC1).
AB - Deciphering the protein posttranslational modification (PTM) code is one of the greatest biochemical challenges of our time. Phosphorylation and ubiquitylation are key PTMs that dictate protein function, recognition, sub-cellular localization, stability, turnover and fate. Hence, failures in their regulation leads to various disease. Chemical protein synthesis allows preparation of ubiquitinated and phosphorylated proteins to study their biochemical properties in great detail. However, monitoring these modifications in intact cells or in cell extracts mostly depends on antibodies, which often have off-target binding. Here, we report that the most widely used antibody for ubiquitin (Ub) phosphorylated at serine 65 (pUb) has significant off-targets that appear during mitosis. These off-targets are connected to polo-like kinase 1 (PLK1) mediated phosphorylation of cell cycle-related proteins and the anaphase promoting complex subunit 1 (APC1).
KW - antibody off-targets
KW - cell cycle
KW - posttranslational modifications
UR - http://www.scopus.com/inward/record.url?scp=85136340836&partnerID=8YFLogxK
U2 - 10.3390/molecules27154867
DO - 10.3390/molecules27154867
M3 - Article
C2 - 35956818
AN - SCOPUS:85136340836
SN - 1420-3049
VL - 27
JO - Molecules
JF - Molecules
IS - 15
M1 - 4867
ER -