Cloning and Expression inEscherichia coliof Full-Length Complementary DNA Coding for Human α1-Antitrypsin

Abstract

A cDNA library prepared from human liver was screened for α₁-antitrypsin, a major constituent of plasma which functions as inhibitor of proteolytic enzymes. The library was screened using a 12-base-long synthetic oligodeoxyribonucleotide corresponding to a known DNA fragment of human α₁-antitrypsin and by hybrid-selection of α₁-antitrypsin mRNA. A plasmid, pULB1523, was identified carrying a cDNA insert of about 1400 bp coding for human α₁-antitrypsin. Restriction mapping and DNA sequence analysis indicated that the 1400 bp code for the signal peptide and for the complete mature α₁-antitrypsin molecule. In addition, a solid-phase enzyme-linked immunoassay showed that pULB1523 expresses human α₁-antitrypsin in bacteria. Fusion of the α₁-antitrypsin sequence to the leader sequence of the β-lactamase gene (plasmid pKT287) resulted also in the expression of the protein in bacteria.