What are the common vector/plasmids used in the Pichia pastoris Protein expression system?
The expression of any recombinant gene in P. pastoris has three phase: (a) Cloning of a new gene into a suitable expression vector, (b) insertion of the cloned vector into P. pastoris host genome; and (c) trial of the potential different strains for the expression of the recombinant integrated gene.
What are the major components in the expression vector of P. pastoris?
The expression vectors in the P. pastoris expression system are one of the major components of this system. These vectors are composed of three sequences:
- Promoter sequence (most often AOX1) in 5′ region
- Transcriptional termination sequence in 3′ region which is essential in the processing and polyadenylation of messenger RNAs
- Single or multiple cloning sites, necessary for the insertion of the gene of interest.
Why expression vector of Pichia pastoris need to be linearized?
The episomal vectors can replicate either autonomously in the cytoplasm or as part of a chromosome. But the vectors used in P. pastoris have no stable episomal status; therefore, they should first be linearized with enzymes and then be integrated into the P. pastoris chromosome.
Common plasmids used in the P. pastoris expression system to produce extracellular and intracellular proteins are listed in Tables 1 and 2, respectively.
| Vector name | Marker gene | Used strain | Recombinant protein |
|---|---|---|---|
| pPIC3.5K | His4, Kan, Amp | KM71 | Maltooligosyltrehalose synthase |
| SMD1168 | Camellia sinensisÊheat shock protein | ||
| GS115 | Pleurotus ostreatusÊlaccases | ||
| GS115 | Rhizopus oryzaeÊLipase | ||
| GS115 | HSA/GH fusion protein | ||
| pPICZ | Shble | X_33 | Aquaporin |
| KM71 | Membrane protein | ||
| KM71 | Dengue virus envelope glycoprotein | ||
| pHIL_D2 | His4, Amp | GS115 | Prostaglandin H synthase_2 |
| GS115 | CatA1 and SODC | ||
| KM71 | RhodococcusÊnitrile hydratase |
Table 1. Common Pichia pastoris expression vectors for the production of intracellular proteins
| Vector name | Marker gene | Used strain | Recombinant protein |
|---|---|---|---|
| pPIC9K | His4, Kan, Amp | GS115 | Xylanase |
| GS115 | Porcine circovirus type 2 | ||
| GS115 | Endo_1,3(4)_b_d_glucanase | ||
| GS115 | Staphylokinase | ||
| pPICZ_ | Shble | SMD1168 | Human chitinase |
| GS115 | Human topoisomerase I | ||
| GS115 | Human interferon gamma | ||
| X_33 | C_reactive protein | ||
| SuperMan5 | Insulin | ||
| X_33 | Human RNase4 | ||
| pHIL_S1 | His4, Amp | GS115 | Rabies virus glycoprotein |
| GS115 | Rhizopus oryzaeÊLipase | ||
| KM71 | Camel lactoferricin | ||
| pGAPZ_ | Shble | GS115 | Acyl homoserine lactonase |
| SMD1168 | Variable lymphocyte receptor B | ||
| X_33 | Human gastric lipase | ||
| pJL_SX | FLD1, Amp | MS105 | Formaldehyde dehydrogenase |
| pBLHIS_SX | His4, Amp | JC100 | Leukocyte protease inhibitor |
Table 2. Common Pichia pastoris expression vectors for the production of secretory proteins
Ref: Pichia pastoris: A highly successful expression system for optimal synthesis of heterologous proteins