Utilization of Sumo Tag and Freeze-Thawing Method for a High-Level Expression and Solubilization of Recombinant Human Angiotensin-Converting Enzyme 2 (Rhace2) Protein in E. Coli Publisher Pubmed



Mohammadi M¹ ; Taheri RA² ; Bemani P³ ; Hashemzadeh MS² ; Farnoosh G¹ ; Amini R⁴
Source: Protein and Peptide Letters Published:2022

Abstract

Background: SARS-CoV-2 uses angiotensin-converting enzyme 2 (ACE2) as a receptor for entering the host cells. Production of the ACE2 molecule is important because of its potency to use as a blocker and therapeutic agent against SARS-CoV-2 for the prophylaxis and treatment of COVID-19. Objective: The recombinant human ACE2 (rhACE2) is prone to form an inclusion body when expressed in the bacterial cells. Methods: We used the SUMO tag fused to the rhACE2 molecule to increase the expression level and solubility of the fusion protein. Afterward, the freeze-thawing method plus 2 M urea solubilized aggregated proteins. Subsequently, the affinity of solubilized rhACE2 to the receptor binding domain (RBD) of the SARS-CoV-2 spike was assayed by ELISA and SPR methods. Results: SUMO protein succeeded in increasing the expression level but not solubilization of the fusion protein. The freeze-thawing method could solubilize and recover the aggregated fusion proteins significantly. Also, ELISA and SPR assays confirmed the interaction between solubilized rhACE2 and RBD with high affinity. Conclusion: The SUMO tag and freeze-thawing method would be utilized for high-level expression and solubilization of recombinant rhACE2 protein. © 2022 Bentham Science Publishers.