Expression and purification optimization of an N-terminal Pfs230 transmission-blocking vaccine candidate

Publication date: Available online 10 April 2019Source: Protein Expression and PurificationAuthor(s): Shwu-Maan Lee, Jordan Plieskatt, Seetha Krishnan, Monika Raina, Rakeshkumar Harishchandra, C. Richter KingAbstractIn an effort to control and eventually eliminate malaria, the development of transmission-blocking vaccines has long been sought. However, few antigens have been evaluated in clinical trials, often due to limitations in the expression and purification of the antigen in sufficient yield and quality. Pfs230, a surface antigen of gametocytes, has recently advanced to clinical evaluation as a conjugate vaccine using the Pseudomonas aeruginosa exoprotein A carrier protein. Here we continue to build upon prior work of developing a Pfs230 candidate in the baculovirus system, Pfs230C1 (aa 443–731), through systematic process development efforts to improve yield and purity.Various insect cells including High Five, Sf9 and Super Sf9 were first evaluated for quality and quantity of antigen, along with three insect cell media. In the selection of Sf9 cells, an intact Pfs230C1 was expressed and harvested at 48 h for downstream development. A downstream process, utilizing immobilized metal affinity column (IMAC), followed by ion exchange (IEX) membranes (Mustang S) and finally IEX chromatography (DEAE) yielded a pure Pfs230C1 protein. The complete process was repeated three times at the 20 L scale.To support the eventual chemistry manufacturing and controls (CMC) of Pfs23...
Source: Protein Expression and Purification - Category: Biochemistry Source Type: research