Part:BBa_K5249034
AOXm-α signal peptid combination of promoters and signaling peptides
The predominant promoter and signal peptide (SP) combination for protein expression in P. pastoris is typically PAOX1 and the α-mating factor (α-MF) prepro-leader from S. cerevisiae. However, it cannot be definitively asserted as the optimal expression pairing for all proteins. Here we chose 3 promoters including AOX1, AOXm and GAP and 2 signal peptides including α and MF. AOX1is transcribed only in response to methanol and is repressed by glucose and ethanol. The transcription level of AOX1 in methanol-grown cells is more than 1000-fold higher than that of glucose-grown cells. The expression of PAOX1 can be regulated with both repression and derepression mechanisms responding to different carbon sources. Many studies of optimizing AOX1 promoter have focused on deletion and duplication of putative transcription factor-binding sites and identification of upstream activation sequence. PAOXm, an enhanced variant, results from deleting the −777 to −721 regions of PAOX1 and contributing a copy of the cis-acting elements from the PAOX1's −230 to −190 regions. This modification augments the expression of foreign proteins in P. pastoris. Glyceraldehydes-3-phosphate dehydrogenase (GAP) promoter has recently been developed for P. pastoris as an alternative strong constitutive promoter [7]. In GAP promoter expression system, the cloned heterologous protein will be expressed along with cell growth if the protein is not toxic to the cell. Therefore, it can easily adapt to a continuous culture that theoretically allows indefinite production of recombinant proteins.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal XhoI site found at 1123
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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