Abstract

Mammalian cells play a key role in the production of protein biopharmaceuticals. However, the protein yield from these cells is often hindered by a bottleneck in the saturated secretory pathway, where a sophisticated mechanism of vesicle trafficking is mediated by numerous proteins, among which are the conserved SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) proteins. SNAREs regulate all the intracellular trafficking pathways and trigger the fusion of transport vesicles with the target membranes. Some of the best characterised mammalian SNAREs include Synaptosome-associated protein of 23 kDa/25 kDa (Snap23/Snap25) and Vesicle associated membrane protein 2/8 (Vamp2/8). The ectopic expression of exocytic SNAREs differentially impacts the secretory capacity of mammalian cells thereby, boosting the secretion of recombinant proteins. Here, we analyzed the effect of different SNAREs on the expression of recombinant secreted proteins. Different reporter plasmids were co-transfected with either single (Snap 23 or Snap 25 or Vamp 8) or multiple (Snap 23+Snap 25 or Snap 23+Vamp 8 or Snap 25+Vamp 8 or Snap 23+Snap 25+Vamp8) SNAREs and the expression level of the reporter proteins was quantified using HiBiT assay. The optimal conditions for co-transfections with one or multiple effector plasmids were determined by testing various effector/reporter plasmid ratios. Overall, the expression of reporter proteins was enhanced by different combinations of SNAREs. Therefore, detailed understanding of SNARE impact on the secretion of heterologous proteins in mammalian cells may foster advances in biopharmaceutical manufacturing.