Enhanced recovery of a secreted recombinant human growth factor using stabilizing additives and by co-expression of human serum albumin in the moss Physcomitrella patens

摘要

The production of pharmaceutical proteins in plants provides a valuable alternative to other traditional eukaryotic expression systems from economic and safety perspectives. The moss Physcomitrella patens allows the expression and secretion of complextarget proteins into a simple aqueous maintenance medium, which facilitates downstream processing by rendering it less complex. To address the question of whether the addition of protein-stabilizing substances enhances the recovery of a target protein secreted into the culture medium, several additives at different concentrations were tested in a small-scale screening system. Although polyvinylpyrrolidone (PVP) and human serum albumin (HSA) showed a significant impact on protein levels, supplementationof the medium with these substances was accompanied by certain limitations in upstream processes, such as foam formation (HSA), and in downstream processes, such as reduced binding efficiency on chromatography columns (PVP), respectively. In order to reap the benefit of the enhancing effect and to avoid the given negative aspects, we developed a new strategy based on the recombinant expression of HSA in plants that are already capable of expressing a target protein. First, we analysed the expression and secretion of recombinant HSA in transiently and stably transformed wild-type (WT) plants. HSA was then co-expressed in Physcomitrella plants transgenic for human vascularendothelial growth, actor(VEGF). Even with high expression levels of recombinant human VEGF (rhVEGF), the' co-expression of recombinant HSA (rHSA) resulted in 48%-102% higher recovery of the target protein without concomitant negative effects on the upstream process. This strategy enables the enhanced recovery of target protein and does not require the addition of foreign components directly to the culture medium.