Work

A Protein Editing Method to Modify Proteins in Live Mammalian Cells

Year: 2026

Type: article

Abstract: Protein editing, as mediated by protein trans-splicing, offers the ability to manipulate, label, and modify proteins in vitro and in live mammalian cells. This ability to observe and control proteins with this high resolution is a burgeoning area of interest within the chemical biology field. In this protocol, we outline the general approach for setting up a cellular protein editing experiment, emphasizing the planning process for a successful protein editing experiment. Additionally, we include methods to install and subsequently label a click chemistry handle p-azido-phenylalanine, a non-canonical amino acid (ncAA) within the intein donor, such that this ncAA and any label can be edited into a protein in live mammalian cells. Overall, we anticipate that this protocol should serve as a generalizable roadmap for planning and carrying out a protein editing experiment. © 2026 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Expression of intein donor containing pAzF Basic Protocol 2: Purification and labeling of recombinant intein donor Basic Protocol 3: Preparation of mammalian cells for editing experiments Basic Protocol 4: Protein editing via electroporation.

Source: Current Protocols

DOI: 10.1002/cpz1.70443

Authors: Jenna N. Beyer, Kaitlyn Toy, George M. Burslem

Institution: University of Pennsylvania

Language: English

Topic: Biochemical and Structural Characterization

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: hybrid

Estimated APC paid: $4,890

Funders: National Science Foundation, National Institutes of Health, National Institute of General Medical Sciences