Kiefer Anna Maria, Niemeyer Justus, Probst Anna, Erkel Gerhard, Schroda Michael
Molecular Biotechnology & Systems Biology, TU Kaiserslautern, Kaiserslautern, Germany.
Front Plant Sci. 2022 Sep 20;13:988870. doi: 10.3389/fpls.2022.988870. eCollection 2022.
The spike protein is the major protein on the surface of coronaviruses. It is therefore the prominent target of neutralizing antibodies and consequently the antigen of all currently admitted vaccines against SARS-CoV-2. Since it is a 1,273-amino acids glycoprotein with 22 N-linked glycans, the production of functional, full-length spike protein was limited to higher eukaryotes. Here we report the production of full-length SARS-CoV-2 spike protein - lacking the C-terminal membrane anchor - as a secreted protein in the prefusion-stabilized conformation in the unicellular green alga . We show that the spike protein is efficiently cleaved at the furin cleavage site during synthesis in the alga and that cleavage is abolished upon mutation of the multi-basic cleavage site. We could enrich the spike protein from culture medium by ammonium sulfate precipitation and demonstrate its functionality based on its interaction with recombinant ACE2 and ACE2 expressed on human 293T cells. is a GRAS organism that can be cultivated at low cost in simple media at a large scale, making it an attractive production platform for recombinant spike protein and other biopharmaceuticals in low-income countries.
刺突蛋白是冠状病毒表面的主要蛋白。因此，它是中和抗体的主要靶点，也是目前所有获批的抗SARS-CoV-2疫苗的抗原。由于它是一种含有22个N-连接聚糖的1273个氨基酸的糖蛋白，功能性全长刺突蛋白的生产仅限于高等真核生物。在此，我们报道了在单细胞绿藻中以分泌蛋白的形式生产处于融合前稳定构象的全长SARS-CoV-2刺突蛋白（缺少C末端膜锚定区）。我们表明，刺突蛋白在绿藻合成过程中在弗林蛋白酶切割位点被有效切割，并且在多碱性切割位点突变后切割被消除。我们可以通过硫酸铵沉淀从培养基中富集刺突蛋白，并基于其与重组ACE2以及在人293T细胞上表达的ACE2的相互作用证明其功能。绿藻是一种公认安全的生物，可以在简单培养基中低成本大规模培养，使其成为低收入国家生产重组刺突蛋白和其他生物制药的有吸引力的平台。
