Abstract
The continuing emergence of variants of the SARS-CoV-2 virus requires the development of modular molecular therapies. Here, we engineered a recombinant amphiphilic protein, oleosin, to spontaneously self-assemble into multivalent micellar nanostructures which can block the Spike S1 protein of SARS-CoV-2 pseudoviruses (PVs). Short recombinant proteins like oleosin can be formulated more easily than antibodies and can be functionalized with precision through genetic engineering. We cloned S1-binding mini-protein genes called LCB x, previously designed by David Baker’s laboratory (UW Seattle), to the N-terminus of oleosin, expressing Oleo-LCB x proteins in E. coli . These proteins largely formed 10-100 nm micelles as verified by dynamic light scattering. Two proteins, Oleo-LCB1 and Oleo-LCB3, were seen to completely and irreversibly block transduction by both wild-type and delta variant PVs into 293T-hsACE2 cells at 10 μM. Presented in multivalent micelles, these proteins reduced transduction by PVs down to a functional protein concentration of 5 nM. Additionally, Oleo-LCB1 micelles outperformed corresponding synthetic LCB1 mini-proteins in reducing transduction by PVs. Tunable aqueous solubility of recombinant oleosin allowed incorporation of peptides/mini-proteins at high concentrations within micelles, thus enhancing drug loading. To validate the potential multifunctionality of the micelles, we showed that certain combinations of Oleo-LCB1 and Oleo-LCB3 performed much better than the individual proteins at the same concentration. These micelles, which we showed to be non-toxic to human cells, are thus a promising step toward the design of modular, multifunctional therapeutics that could bind to and inactivate multiple receptors and proteins necessary for the infection of the SARS-CoV-2 virus.
Keywords: Delta Variant; Mini-Proteins; Multivalent Micelles; Oleosin; Peptides; Self-Assembly.
【저자키워드】 delta variant, peptides, Self-Assembly., Oleosin, Multivalent Micelles, Mini-Proteins, 【초록키워드】 antibody, Genetic, variant, Infection, Delta, Proteins, Laboratory, Protein, multivalent, receptor, molecular, Therapies, Combination, Concentration, Recombinant protein, Precision, E. coli, human cells, wild-type, S1 protein, DAVID, functional protein, Seattle, N-terminus, Cell, non-toxic, performed, cloned, reduced, reducing, expressing, multifunctional, the Spike, Baker, SARS-CoV-2 pseudovirus, scattering, the SARS-CoV-2 virus, 【제목키워드】 SARS-CoV-2, transduction, recombinant, block, micelle,