Molecular basis of αl-antitrypsin deficiency and emphysema associated with the αl-antitrypsin M_{mineral springs} allele

The M_{mineral springs} α1-antitrypsin (α1AT) allele, causing α1AT deficiency and emphysema, is unique among the α1AT-deficiency alleles in that it was observed in a black family, whereas most mutations causing α1AT deficiency are confined to Caucasian populations of European descent. Immobilized pH gradient analysis of serum demonstrated that α1AT M_{mineral springs} migrated cathodal to the normal M2 allele. Evaluation of M_{mineral springs} α1AT as an inhibitor of neutrophil elastase, its natural substrate, demonstrated markedly lower than normal function. Characterization of the α1AT M_{mineral springs} gene demonstrated that it differed from the common normal M1(Ala²¹³) allele by a single-base substitution causing the amino acid substitution Gly-67 (GGG)→Glu-67 (GAG). Capitalizing on the fact that this mutation creates a polymorphism for the restriction endonuclease AvaII, family analysis demonstrated that the M_{mineral springs} α1AT allele was transmitted in an autosomal-codominant fashion. Evaluation of genomic DNA showed that the index case was homozygous for the α1ATM_{mineral springs} allele. Cytoplasmic blot analysis of blood monocytes of the M_{mineral springs} homozygote demonstrated levels of α1AT mRNA transcripts comparable to those in cells of a normal M1(Val²¹³) homozygote control. Evaluation of in vitro translation of M_{mineral springs} α1AT mRNA transcripts demonstrated a normal capacity to direct the translation of αAT. Evaluation of secretion of α1AT by the blood monocytes by pulse-chase labeling with [³⁵S]methionine, however, demonstrated less secretion by the M_{mineral springs} cells than normal cells. To characterize the posttranslational events causing the α1AT-secretory defect associated with the α1AT M_{mineral springs} gene, retroviral gene transfer was used to establish polyclonal populations of murine fibroblasts containing either a normal human M1 α1AT cDNA or an M_{mineral springs} α1AT cDNA and expressing comparable levels of human α1AT mRNA transcripts. Pulse-chase labeling of these cells with [³⁵S]methionine demonstrated less secretion of human α1AT from the M_{mineral springs} cells than from the M1 cells, and evaluation of cell lysates also demonstrated lower amounts of intracellular human α1AT in the M_{mineral springs} cells tnan in the normal M1 control cells. Thus, the Gly-67→Glu mutation that characterizes M_{mineral springs} causes reduced α1AT secretion on the basis of aberrant posttranslational α1AT biosynthesis by a mechanism distinct from that associated with the α1AT Z allele, whereby intracellular aggregation of the mutant protein is etiologic of the α1AT-secretory defect. Furthermore, for the α1AT protein that does reach the circulation, this mutation markedly affects the ability of the molecule to inhibit neutrophil elastase; i.e., the α1AT M_{mineral springs} allele predisposes to emphysema on the basis of serum α1AT deficiency coupled with α1AT dysfunction.