Identification of alpha(1)-antitrypsin variants in plasma with the use of proteomic technology

Background: Proteomic technology permits the investigation of genetic metab olic diseases at the level of protein expression. Changes in the expression , polypeptide structure, and posttranslational modification of individual p roteins can be detected in complex mixtures of proteins. Methods: We used high-resolution two-dimensional polyacrylamide gel electro phoresis to separate isoforms of plasma proteins and detect abnormalities o f mass and/or charge. We confirmed the identity of the separated proteins b y in-gel digestion with proteases and N-glycanases and then analyzed the re leased peptides and glycans by matrix-assisted laser-desorption ionization- time-of-flight mass spectrometry. Results: Complete characterization of the polypeptide sequences and glycosy lation of alpha (1)-antitrypsin isoforms was achieved in plasma from contro ls and from patients with three different known alpha (1)-antitrypsin defic iencies and congenital disorder of glycosylation type Ia. Conclusions: This study shows that proteomic techniques are a powerful and sensitive means of detecting changes in the amino acid sequence and abnorma l posttranslational modifications of specific proteins in a complex biologi c matrix. (C) 2001 American Association for Clinical Chemistry.