Recombinant expression, purification and PEGylation of Paneth cell peptide (cryptdin-2) with value added attributes against Staphylococcus aureus.

Scientific Reports
Navneet KaurPraveen Rishi

Abstract

Cryptdins are disulfide-rich cationic antimicrobial peptides secreted by mouse Paneth cells and are known to exhibit potent antimicrobial activity against various deadly pathogens. Keeping in view the extremely low yield obtained from mouse Paneth cells and high cost of synthetic peptide(s), herein, we have attempted to produce cryptdin-2 in Escherichia coli using recombinant technology. To avoid lethal effects of peptide on the host cells, cryptdin-2 was expressed as a fusion protein with thioredoxin as fusion partner which yielded 40 mg/L protein in the soluble fraction. Subsequently, mature cryptdin-2 was cleaved from the fusion partner and purified by cation exchange chromatography. Since conjugation of poly(ethylene) glycol (PEG) has been known to improve the biological properties of biomolecules, therefore, we further attempted to prepare PEG-conjugated variant of cryptdin-2 using thiol specific PEGylation. Though the antimicrobial activity of PEGylated cryptdin-2 was compromised to some extent, but it was found to have enhanced serum stability for longer duration as compared to its un-modified forms. Also, it was found to exhibit reduced toxicity to the host cells. Further, its synergism with gentamicin suggests that PEG...Continue Reading

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Citations

Feb 9, 2021·Journal of Peptide Science : an Official Publication of the European Peptide Society·Taoran WangChenhong Wang
Dec 16, 2020·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Verónica RojasFanny Guzmán
Jun 8, 2021·Biomaterials Science·Zixian CuiRachel A Letteri

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Methods Mentioned

BETA
gel filtration
Scanning Electron Microscopy
Fluorescence
gel-filtration
flow cytometry
FACS

Software Mentioned

FlowJo
Emini Accessibility

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