Scalable Production of Recombinant Three-finger Proteins: from Inclusion Bodies to High Quality Molecular Probes

Overview

Journal Microb Cell Fact

Publisher Biomed Central

Specialties Biotechnology
Microbiology

Date 2024 Feb 12

PMID 38347541

Authors

Jiang Xu

Xiao Lei

Ao Li

Jun Li

Shuxing Li

Lin Chen

Affiliations

Soon will be listed here.

Abstract

Background: The three-finger proteins are a collection of disulfide bond rich proteins of great biomedical interests. Scalable recombinant expression and purification of bioactive three-finger proteins is quite difficult.

Results: We introduce a working pipeline for expression, purification and validation of disulfide-bond rich three-finger proteins using E. coli as the expression host. With this pipeline, we have successfully obtained highly purified and bioactive recombinant α-Βungarotoxin, k-Bungarotoxin, Hannalgesin, Mambalgin-1, α-Cobratoxin, MTα, Slurp1, Pate B etc. Milligrams to hundreds of milligrams of recombinant three finger proteins were obtained within weeks in the lab. The recombinant proteins showed specificity in binding assay and six of them were crystallized and structurally validated using X-ray diffraction protein crystallography.

Conclusions: Our pipeline allows refolding and purifying recombinant three finger proteins under optimized conditions and can be scaled up for massive production of three finger proteins. As many three finger proteins have attractive therapeutic or research interests and due to the extremely high quality of the recombinant three finger proteins we obtained, our method provides a competitive alternative to either their native counterparts or chemically synthetic ones and should facilitate related research and applications.

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