A Dual-reporter System for Investigating and Optimizing Protein Translation and Folding in E. Coli

Overview

Journal Nat Commun

Specialty Biology

Date 2021 Oct 20

PMID 34667164

Citations 11

Authors

Ariane Zutz

Louise Hamborg

Lasse Ebdrup Pedersen

Maher M Kassem

Elena Papaleo

Anna Koza

Markus J Herrgard

Sheila Ingemann Jensen

Kaare Teilum

Kresten Lindorff-Larsen

Alex Toftgaard Nielsen

Affiliations

Soon will be listed here.

Abstract

Strategies for investigating and optimizing the expression and folding of proteins for biotechnological and pharmaceutical purposes are in high demand. Here, we describe a dual-reporter biosensor system that simultaneously assesses in vivo protein translation and protein folding, thereby enabling rapid screening of mutant libraries. We have validated the dual-reporter system on five different proteins and find an excellent correlation between reporter signals and the levels of protein expression and solubility of the proteins. We further demonstrate the applicability of the dual-reporter system as a screening assay for deep mutational scanning experiments. The system enables high throughput selection of protein variants with high expression levels and altered protein stability. Next generation sequencing analysis of the resulting libraries of protein variants show a good correlation between computationally predicted and experimentally determined protein stabilities. We furthermore show that the mutational experimental data obtained using this system may be useful for protein structure calculations.

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