Optimized protocol for soluble prokaryotic expression, purification and refolding of the human inhibin α subunit, a cysteine rich peptide chain

Kolivand, Sedighe; Nazari, Mahboobeh; Modarressi, Mohammad Hossein; Najafabadi, Mohammad Reza H.; Hemati, Atefeh; Ghafouri‐Fard, Soudeh; Motevaseli, Elahe

doi:10.3233/hab-190399

Cited by 1 publication

(1 citation statement)

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“…However, the presence of cysteine residues may have an adverse effect on the expression of soluble proteins in E. coli or the refolding of denatured proteins. − This was the situation we found for apoptin. Sometimes, the C-terminal cysteine has to be removed to stabilize the protein and facilitate purification.…”

Section: Discussionmentioning

confidence: 65%

Highly Efficient Method for Intracellular Delivery of Proteins Mediated by Cholera Toxin-Induced Protein Internalization

et al. 2021

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Delivery of functional proteins into cells may help us understand how specific protein influences cell behavior as well as treat diseases caused by protein deficiency or loss-of-function mutations. However, protein cannot enter cells by diffusion. In this work, a novel cell biology tool for delivering recombinant proteins into mammalian cells was developed. We hijacked the intracellular transport routes used by the cholera toxin and took advantage of recent development on split intein that is compatible with denatured conditions and shows an exceptional splicing activity to deliver a protein of interest into mammalian cells. Here, we used green fluorescent protein and apoptin as proofs-of-concept. The results demonstrate that the cholera toxin B subunit alone could deliver other recombinant proteins into cells through either covalent conjugation or noncovalent interaction. Our method offers more than 10-fold better delivery efficiency than the tat cellpenetrating peptide and is selective for ganglioside-rich cells. This study adds a useful tool to the receptor-mediated intracellular targeting toolkit and opens possibility for the selective delivery of therapeutic proteins into ganglioside-rich cells.

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