Simultaneous quantification of total and conjugated ubiquitin levels in a single immunoblot

Choongseob Oh; Jong Hyuk Yoon; Soonyong Park; Yung Joon Yoo

doi:10.1016/j.ab.2013.09.011

Simultaneous quantification of total and conjugated ubiquitin levels in a single immunoblot

Anal Biochem. 2013 Dec 15;443(2):153-5. doi: 10.1016/j.ab.2013.09.011. Epub 2013 Sep 16.

Authors

Choongseob Oh¹, Jong Hyuk Yoon, Soonyong Park, Yung Joon Yoo

Affiliation

¹ School of Life Sciences, Gwangju Institute of Science and Technology, Gwangju 500-712, Republic of Korea.

PMID: 24050967
DOI: 10.1016/j.ab.2013.09.011

Abstract

Ubiquitin (Ub) is a posttranslational modifier, and total Ub (UbT) is always in dynamic equilibrium among free Ub (UbF), activated Ub (UbA), and conjugated Ub (UbC) in the forms of mono-Ub, thioester-bond-linked Ub, and peptide-bond-linked Ub, respectively. In this study, we developed a simple method to simultaneously determine the levels of UbT, UbF+UbA, and UbC in a single immunoblot and demonstrated its reliability and reproducibility by determining [UbT], [UbF+UbA], and [UbC] in various mouse tissues and cultured cells.

Keywords: Ubiquitin; Ubiquitination; Western blot.

Publication types

Evaluation Study
Research Support, Non-U.S. Gov't

MeSH terms

Animals
Blotting, Western / methods*
Cell Line
Humans
Mice
Peptides / analysis
Reproducibility of Results
Sulfides / analysis
Ubiquitin / analysis*

Substances

Peptides
Sulfides
Ubiquitin