Probst Anna, Knochenhauer Doreen, Niemeyer Justus, Fischer Laura, Schroda Michael
Molecular Biotechnology & Systems Biology, RPTU Kaiserslautern-Landau, Paul- Ehrlich-Straße 23, D-67663, Kaiserslautern, Germany.
Curr Genet. 2025 Mar 19;71(1):7. doi: 10.1007/s00294-025-01311-2.
There is great interest in establishing microalgae as new platforms for the sustainable production of high-value products such as recombinant proteins. Many human therapeutic proteins must be glycosylated, which requires their passage through the secretory pathway into the culture medium. While the low complexity of proteins in the culture medium should facilitate affinity purification of secreted recombinant proteins, this has proven challenging for proteins secreted by the unicellular green alga Chlamydomonas reinhardtii. In Leishmania tarentulae, we observed that C-terminally exposed affinity tags are frequently truncated, presumably due to proteolytic activity. We wondered whether this might also occur in Chlamydomonas and contribute to the difficulties in affinity purification of secreted proteins in this alga. Using the methionine-rich 2S albumin from Bertholletia excelsa and the ectodomain of the SARS-CoV-2 spike protein produced and secreted in Chlamydomonas, we demonstrate that they can be efficiently affinity-purified from the culture medium by Ni-NTA chromatography when the 8xHis affinity tag is internalized. This finding represents an important step towards further development of Chlamydomonas as a host for the sustainable production of high-value recombinant proteins.
人们对将微藻建立为生产重组蛋白等高价值产品的可持续新平台有着浓厚兴趣。许多人类治疗性蛋白质必须进行糖基化，这需要它们通过分泌途径进入培养基。虽然培养基中蛋白质的低复杂性应有助于分泌型重组蛋白的亲和纯化，但事实证明，这对于单细胞绿藻莱茵衣藻分泌的蛋白质来说具有挑战性。在塔兰托利什曼原虫中，我们观察到C端暴露的亲和标签经常被截断，推测是由于蛋白水解活性。我们想知道这种情况在衣藻中是否也会发生，并导致这种藻类中分泌蛋白亲和纯化的困难。利用衣藻中产生和分泌的巴西栗富含蛋氨酸的2S白蛋白以及严重急性呼吸综合征冠状病毒2（SARS-CoV-2）刺突蛋白的胞外结构域，我们证明，当8xHis亲和标签内化时，可以通过镍-亚氨基二乙酸（Ni-NTA）色谱法从培养基中高效地亲和纯化它们。这一发现是将衣藻进一步开发成为可持续生产高价值重组蛋白宿主的重要一步。
