Expression and Purification of Membrane Proteins in Different Hosts

Kahaki, Fatemeh Abarghooi; Monzavi, Sakineh; Bamehr, Hadi; Bandani, Eshagh; Payandeh, Zahra; Jahangiri, Abolfazl; Khalili, Saeed

doi:10.1007/s10989-019-10009-2

Cited by 4 publications

(1 citation statement)

References 120 publications

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“…However, the biological activity of this recombinant protein is not demonstrated by the vendor. Since the proper posttranslational modifications such as the formation of disulfide bonds, glycosylation, and phosphorylation are not warranted in the cell‐free wheat germ protein expression system, 43 this cell‐free protein expression system does not appear to be ideal for the production of PDGF D dimer species which require proper posttranslational modifications. In the current study, using the second generation MVA virus protein expression system combined with high performance HisTrap IMAC, we successfully produced and purified recombinant PDGF D dimer species which main biological activity and chemical integrity.…”

Section: Discussionmentioning

confidence: 99%

The production of recombinant platelet‐derived growth factor D using the second generation modified vaccinia Ankara viral system

et al. 2022

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The vaccinia virus expression system is known for the efficient production of recombinant proteins with “appropriate” posttranslational modification using desired mammalian cell lines. However, being a replication competent virus, vaccinia virus poses a health threat to immunocompromised individuals and requires biosafety level 2 (BSL2) laboratory precautions, thereby restricting its use by the scientific community. Development of the host range restricted modified vaccinia Ankara (MVA) system has allowed researchers to work with a safer virus even at BSL1. Here, we report on the use of an improved second generation MVA viral system incorporating two selective markers and fluorescent proteins for easier recombinant virus identification. Notably, we demonstrate that this novel system is capable of producing secreted recombinant proteins, a finding not previously reported. Through purification and characterization of wild type and mutant platelet‐derived growth factor D (PDGF D) dimer species, we demonstrate this system is capable of producing the latent full‐length PDGF D dimer, partially processed intermediate dimer (hemidimer), as well as fully processed growth factor domain dimer that show chemical integrity and biological activity. Importantly, this system is amenable to scaling up for the mass production of recombinant PDGF D (rPDGF D) dimer species.

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Section: Discussionmentioning

confidence: 99%

The production of recombinant platelet‐derived growth factor D using the second generation modified vaccinia Ankara viral system

et al. 2022

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A unique antigen against SARS-CoV-2, Acinetobacter baumannii, and Pseudomonas aeruginosa

Rahbar

Mubarak

Hessami

et al. 2022

Sci Rep

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The recent outbreak of COVID-19 has increased hospital admissions, which could elevate the risk of nosocomial infections, such as A. baumannii and P. aeruginosa infections. Although effective vaccines have been developed against SARS-CoV-2, no approved treatment option is still available against antimicrobial-resistant strains of A. baumannii and P. aeruginosa. In the current study, an all-in-one antigen was designed based on an innovative, state-of-the-art strategy. In this regard, experimentally validated linear epitopes of spike protein (SARS-CoV-2), OmpA (A. baumannii), and OprF (P. aeruginosa) were selected to be harbored by mature OmpA as a scaffold. The selected epitopes were used to replace the loops and turns of the barrel domain in OmpA; OprF311–341 replaced the most similar sequence within the OmpA, and three validated epitopes of OmpA were retained intact. The obtained antigen encompasses five antigenic peptides of spike protein, which are involved in SARS-CoV-2 pathogenicity. One of these epitopes, viz. QTQTNSPRRARSV could trigger antibodies preventing super-antigenic characteristics of spike and alleviating probable autoimmune responses. The designed antigen could raise antibodies neutralizing emerging variants of SARS-CoV-2 since at least two epitopes are consensus. In conclusion, the designed antigen is expected to raise protective antibodies against SARS-CoV-2, A. baumannii, and P. aeruginosa.

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Challenges and Solutions in the Recombinant Expression of Membrane Proteins

Liu,

He,

Tian

et al. 2023

PPL

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Membrane proteins are important components of the proteome and play key roles in many biological processes, such as signal transduction, material transport, cell recognition, etc. Membrane proteins are involved in several fields, and more and more researchers want to understand them. However, the structural properties of membrane proteins make their recombinant expression yield low. This adversely affects the study of the structure and function of membrane proteins. Therefore, it is crucial to have a comprehensive and up-to-date understanding of membrane protein recombinant expression. Based on the current stage of research on membrane proteins, the article describes the current challenges faced by membrane protein recombinant expression and the solutions that can be applied to lay the foundation for a better study of membrane proteins in the future.

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Expression and Purification of Membrane Proteins in Different Hosts

Cited by 4 publications

References 120 publications

The production of recombinant platelet‐derived growth factor D using the second generation modified vaccinia Ankara viral system

The production of recombinant platelet‐derived growth factor D using the second generation modified vaccinia Ankara viral system

A unique antigen against SARS-CoV-2, Acinetobacter baumannii, and Pseudomonas aeruginosa

Challenges and Solutions in the Recombinant Expression of Membrane Proteins

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