Knapp Michael, Kohl Vanessa, Best Tatjana, Rammo Oliver, Ebert Sybille
Faculty of Natural Sciences, Ulm University, Ulm, Germany.
Institute of Applied Biotechnology, Biberach University of Applied Sciences, Biberach, Germany.
Front Bioeng Biotechnol. 2024 Jan 17;11:1319916. doi: 10.3389/fbioe.2023.1319916. eCollection 2023.
The current trend in biopharmaceutical drug manufacturing is towards increasing potency and complexity of products such as peptide scaffolds, oligonucleotides and many more. Therefore, a universal affinity purification step is important in order to meet the requirements for cost and time efficient drug production. By using a self-splicing intein affinity tag, a purification template is generated that allows for a universal chromatographic affinity capture step to generate a tagless target protein without the use of proteases for further tag removal. This study describes the successful implementation of gp41-1-based split inteins in a chromatographic purification process for, e.g., -derived targets. The tagless target is generated in a single-step purification run. The on-column cleavage is induced by triggering a simple pH change in the buffer conditions without the need for additives such as Zn or thiols. This system has proven to be reusable for at least ten purification cycles that use 150 mM HPO as the cleaning agent.
生物制药药物制造的当前趋势是提高产品的效力和复杂性，如肽支架、寡核苷酸等。因此，一个通用的亲和纯化步骤对于满足成本和时间高效的药物生产要求很重要。通过使用自剪接内含肽亲和标签，生成了一个纯化模板，该模板允许进行通用的色谱亲和捕获步骤，以产生无标签的目标蛋白，而无需使用蛋白酶进行进一步的标签去除。本研究描述了基于gp41-1的分裂内含肽在例如衍生目标的色谱纯化过程中的成功应用。无标签目标在单步纯化过程中产生。通过在缓冲条件下触发简单的pH变化来诱导柱上切割，而无需诸如锌或硫醇等添加剂。该系统已被证明至少可重复使用十次纯化循环，使用150 mM HPO作为清洗剂。
