A production platform for disulfide-bonded peptides in the periplasm of Escherichia coli
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View abstract on PubMed
Summary
This summary is machine-generated.This study presents a new method for producing disulfide-bonded peptides in E. coli using the CASPON™ tag, achieving high yields and purity. This recombinant peptide production system offers a sustainable and efficient alternative to chemical synthesis.
Area Of Science
- Biotechnology
- Protein Engineering
- Microbial Fermentation
Background
- Recombinant peptide production in Escherichia coli is a sustainable alternative to chemical synthesis but faces challenges with yield and disulfide bond formation.
- In vivo production of disulfide-bonded peptides is hindered by host proteases/peptidases and the need for periplasmic translocation.
Purpose Of The Study
- To establish an efficient expression system for soluble production of disulfide-bonded peptides in the E. coli periplasm.
- To evaluate the efficacy of the CASPON™ tag in enhancing peptide expression and solubility.
- To investigate peptide degradation pathways and optimize production in industrial settings.
Main Methods
- Expression of model peptides (parathyroid hormone 1-84, somatostatin 1-28, plectasin, aprotinin) with and without the CASPON™ tag in E. coli.
- Utilized BioLector™ cultivations for microliter-scale screening and bioreactor fed-batch cultivations for industrial scale-up.
- Investigated degradation by comparing co- and post-translational signal sequences; confirmed disulfide bond formation via mass spectrometry.
Main Results
- The CASPON™ tag significantly improved the expression of most model peptides, which were otherwise weakly or undetectably expressed.
- Peptide degradation was found to occur predominantly after translocation into the periplasm.
- Achieved high titers (0.6-2.6 g L⁻¹) in industry-relevant strains, exceeding previously published data, with complete expression and disulfide bond formation.
Conclusions
- The CASPON™ technology provides a highly efficient platform for producing soluble disulfide-bonded peptides in the E. coli periplasm.
- This upstream platform offers unprecedented titers for several model peptides, outperforming existing methods and chemical synthesis.
- The developed system demonstrates significant potential for industrial-scale recombinant peptide manufacturing.
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