Cloning and expression in Escherichia coli of full-length complementary DNA coding for human alpha 1-antitrypsin.
A cDNA library prepared from human liver was screened for alpha 1-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 alpha 1-antitrypsin and by hybrid-selection of alpha 1-antitrypsin mRNA. A plasmid, pULB1523, was identified carrying a cDNA insert of about 1400 bp coding for human alpha 1-antitrypsin. Restriction mapping and DNA sequence analysis indicated that the 1400 bp code for the signal peptide and for the complete mature alpha 1-antitrypsin molecule. In addition, a solid-phase enzyme-linked immunoassay showed that pULB1523 expresses human alpha 1-antitrypsin in bacteria. Fusion of the alpha 1-antitrypsin sequence to the leader sequence of the beta-lactamase gene (plasmid pKT287) resulted also in the expression of the protein in bacteria.
D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D004262:DNA Restriction Enzymes; D004269:DNA, Bacterial / Q000235:genetics; D004274:DNA, Recombinant; D004926:Escherichia coli / Q000235:genetics*; D006801:Humans; D010957:Plasmids; D012333:RNA, Messenger / Q000235:genetics; D000515:alpha 1-Antitrypsin / Q000235:genetics*
D004269:DNA, Bacterial; D004274:DNA, Recombinant; D012333:RNA, Messenger; D000515:alpha 1-Antitrypsin; D004247:DNA; D004262:DNA Restriction Enzymes