Other variants are less functional and are termed A-L and N-Z, dependent on whether they run proximal or distal to the M band. The presence of deviant bands on IEF can signify the presence of alpha-1 antitrypsin deficiency. Since the number of identified mutations has exceeded the number of letters in the alphabet, subscripts have been added to most recent discoveries in this area, as in the Pittsburgh mutation described above. As every person has two copies of the A1AT gene, a heterozygote with two different copies of the gene may have two different bands showing on electrofocusing, although heterozygote with one null mutant that abolishes expression of the gene will only show one band. In blood test results, the IEF results are notated as in PiMM, where Pi stands for protease inhibitor and "MM" is the banding pattern of that patient. Alpha-1 antitrypsin levels in the blood depend on the genotype. Some mutant forms fail to fold properly and are, thus, targeted for destruction in the proteasome, whereas others have a tendency to polymerise, being retained in the endoplasmic reticulum.
Biochemical properties
A1AT is a single-chain glycoprotein consisting of 394 amino acids in the mature form and exhibits many glycoforms. The three N-linked glycosylations sites are mainly equipped with so-called diantennary N-glycans. However, one particular site shows a considerable amount of heterogeneity since tri- and even tetraantennary N-glycans can be attached to the Asparagine 107 (UniProtKB amino acid nomenclature). These glycans carry different amounts of negatively charged sialic acids; this causes the heterogeneity observed on normal A1AT when analysed by isoelectric focusing. Also, the fucosylated triantennary N-glycans were shown to have the fucose as part of a so-called Sialyl Lewis x epitope, which could confer this protein particular protein-cell recognition properties. The single cysteine residue of A1AT in position 256 (UniProtKB nomenclature) is found to be covalently linked to a free single cysteine by a disulfide bridge. Analysis
The level of A1AT in serum is most often determined by adding an antibody that binds to A1AT, then using turbidimetry to measure how much A1AT is present. Other detection methods include the use of enzyme-linked-immuno-sorbent-assays and radial immunodiffusion. Different analytical methods are used to determine A1AT phenotype. As protein electrophoresis is imprecise, the A1AT phenotype is analysed by isoelectric focusing (IEF) in the pH range 4.5-5.5, where the protein migrates in a gel according to its isoelectric point or charge in a pH gradient. Normal A1AT is termed M, as it migrates toward the center of such an IEF gel.