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.