Synthesis in yeast of a functional oxidation-resistant mutant of human alpha-antitrypsin.
Cumulative damage to lung tissue by leukocyte elastase is thought to be responsible for the development of pulmonary emphysema, an irreversible lung disease characterized by loss of lung elasticity. It is also thought to be involved in the rapidly developing and usually fatal adult respiratory distress syndrome. The primary defence against elastase damage is the anti-protease known as alpha 1-antitrypsin, a glycosylated serum protein of 394 amino acids. Oxidation of the methionine 358 residue located at the active centre of alpha 1-antitrypsin results in a dramatic decrease in inhibitory activity towards elastase which effectively inactivates the protective function. It has been suggested that this oxidation sensitivity has a regulatory function and allows tissue breakdown at sites of inflammation by inactivation of alpha 1-antitrypsin by oxygen radicals released by phagocytes. In the above diseases, however, the oxidative inactivation of alpha 1-antitrypsin is probably of major importance in allowing lung damage by elastase. An oxidation-resistant alpha 1-antitrypsin required for emphysemics and provide treatment for acute inflammatory respiratory conditions. To further the possibility of therapy for the above conditions, we describe here the synthesis in yeast of active, non-glycosylated, human alpha 1-antitrypsin. Site-directed mutagenesis has been used to construct an active, oxidation-resistant derivative containing a single methionine to valine substitution at the active centre. This demonstrates the potential of engineered modifications to protein molecules designed to improve their physiological function.
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D005796:Genes; D006801:Humans; D008970:Molecular Weight; D009154:Mutation; D010084:Oxidation-Reduction; D010957:Plasmids; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D000515:alpha 1-Antitrypsin / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
D000515:alpha 1-Antitrypsin; D004247:DNA