JoVE Logo

Sign In

A subscription to JoVE is required to view this content. Sign in or start your free trial.

Abstract

Developmental Biology

Combining Non-reducing SDS-PAGE Analysis and Chemical Crosslinking to Detect Multimeric Complexes Stabilized by Disulfide Linkages in Mammalian Cells in Culture

Published: May 2nd, 2019

DOI:

10.3791/59483

1Department of Molecular Biology, Rowan University, School of Osteopathic Medicine and Graduate School of Biomedical Sciences

Abstract

The structures of many proteins are stabilized through covalent disulfide linkages. In recent work, this bond has also been classified as a post-translational modification. Thus, it is important to be able to study this modification in living cells. A simple method to analyze these cysteine-stabilized multimeric complexes is through a two-step method of non-reducing SDS-PAGE analysis and formaldehyde cross-linking. This two-step method is advantageous as the first step to uncovering multimeric complexes stabilized by disulfide linkages due to its technical ease and low cost of operation. Here, the human bone osteosarcoma cell line U-2 OS is used to illustrate this method by specifically analyzing the nuclear isoform of dUTPase.

Explore More Videos

Keywords Disulfide Linkages

This article has been published

Video Coming Soon

JoVE Logo

Privacy

Terms of Use

Policies

Research

Education

ABOUT JoVE

Copyright © 2024 MyJoVE Corporation. All rights reserved