Expression, purification and characterization of recombinant Z α1-Antitrypsin—The most common cause of α1-Antitrypsin deficiency

α₁-Antitrypsin (α₁AT), the most abundant proteinase inhibitor circulating in the blood, protects extracellular matrix proteins of the lung against proteolytic destruction by neutrophil elastase. α₁AT deficiency predisposes patients to emphysema, juvenile cirrhosis and hepatocellular carcinoma. Over 90% of clinical cases of severe α₁AT deficiency are caused by the Z variant (E342K) of α₁AT. The presence of the Z mutation results in misfolding and polymerization of α₁AT. Due to its inherent propensity to polymerize there are no reported cases of recombinant Z α₁AT production. This has created a major impediment to studying the effect of the Z mutation on α₁AT. Here we report our attempts to produce recombinant Z α₁AT using both Escherichia coli and Pichia pastoris as host systems. Using a range of expression vectors in E. coli we were unable to produce soluble active Z α₁AT. Cytosolic expression of the Z α₁AT gene in P. pastoris was successful. Monomeric and active recombinant Z α₁AT was purified from the yeast cytosol using affinity chromatography and anion exchange chromatography. Biochemical analyses demonstrated that the recombinant Z α₁AT has identical properties to its native counterpart purified from plasma of patients homozygous for the Z allele. A recombinant source of pathological Z α₁AT will increase the chances of elucidating the mechanism of its polymerization and thus the development of therapeutic strategies.