Background: Human alpha 1-antitrypsin (AAT) is a potent inhibitor of multiple serine proteases, and
protects tissues against their harmful effects. Individuals with reduced or abnormal production of this
inhibitor need intravenous administration of exogenous protein. In this study, we employed the
methylotrophic (methanol utilizing) yeast
Pichia pastoris
(
P. pastoris) as a preferential host for efficient
production and secretion of recombinant AAT. Furthermore, we examined different strategies to
maximize the yield of the secreted protein.
Results: Our findings revealed that optimizing the codon usage of AAT gene for
P. pastoris had
positive effects on the level of secreted AAT under the control of inducible alcohol oxidase 1 (AOX1)
and constitutive glycerol aldehyde phosphate dehydrogenase (GAP) promoters. Compared to AOX1,
the GAP promoter increased the yield of AAT by more than two fold. It was also demonstrated that the
human AAT native signal sequence was more effective than the well-known yeast signal sequence,
alpha mating factor (α-MF). Doubling gene dosage nearly doubled the production of AAT, though
dosages exceeding this limit had negative effects on the yield.
Conclusion: P. pastoris is shown to be an efficient expression system for production of recombinant
and biologically active AAT. Also different strategies could be used to elevate the amount of this
secretable protein.
