We are trying to make VLPs in insect cells where the viral protein and the protease that should cleave it are co-expressed from the same virus (both genes cloned into pFastBacDual). The processing is not working well, so now the user would like to try putting the protease under the control of a weaker promoter to lower its expression level.
Does anyone know of a good promoter to use for this purpose? I have read that CMV can be used for this, but this was tried with a similar virus and didn't give good results, so they were interested in trying something different. Any ideas?
maybe the ie1 promoter (+hr5 enhancer) used in the pIEX transient expression construct could be useful. Even if you can use this promoter in transient expression mode for preparative amounts of protein, the copy number of the virus genome at an early stage of virus replication might be much lower than plasmid copy number during transient expression.
It is probably weak enough and early expression of lower amounts of protease could be helpful for processing of the target proteins during synthesis and well-tolerated by the host cells.
Hope this helps.