Molecular analysis of the heterogeneity among the P-family of alpha-1-antitrypsin alleles

M. D. Holmes, M. L. Brantly, Ronald Crystal

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

The rare P-family of α1-antitrypsin (α1AT) variants is defined by the position of migration of the α1AT protein on isoelectric focusing of serum (IEF) between the common M and S variants. To begin to examine the molecular heterogeneity among the P-type alleles, two unrelated subjects and their families identified by IEF to be carrying a P allele were analyzed. The first, P(lowell), is a deficiency allele associated with reduced serum α1AT levels, and the second, P(saint albans), is associated with normal serum levels. DNA sequence analysis of P(lowell), the more anodal of the two variants on IEF analysis, showed that it differed from the normal M1(Val213) allele by a single base and amino acid substitution Asp256 GAT → Val GTT. In contrast, P(saint albans), a slightly more cathodally positioned variant on IEF analysis, differed from the coding exons of the normal M1(Val213) allele by two mutations, Asp341 GAC → Asn AAC, and a silent substitution in the same codon as the P(lowell) variant, Asp256 GAT → Asp GAC. Evaluation of P(lowell) mRNA transcripts by Northern and cytoblot analyses demonstrated they were of normal size and amount, and P(lowell) mRNA transcripts could be translated normally in vitro. Retroviral insertion of the P(lowell) cDNA into the genome of 3T3 fibroblasts demonstrated that it directed the synthesis of α1AT, but at levels 24% that of the P(saint albans) cDNA or the normal M1 (Val213) cDNA, with a pattern of biosynthesis consistent with the concept that the P(lowell) α1AT deficiency state results from intracellular degradation of the newly synthesized P(lowell) protein. In the context that the serum α1AT deficiency associated with other α1AT deficiency mutations resulting from intracellular degradation of α1AT can be overcome by administering estrogenlike drugs, administration of tamoxifen to a subject with the P(lowell)Z phenotype resulted in α1AT serum levels rising 48% over a 5-month period, from below the threshold for protection from emphysema (11 μM) to above that threshold.

Original languageEnglish
Pages (from-to)1185-1192
Number of pages8
JournalAmerican Review of Respiratory Disease
Volume142
Issue number5
DOIs
Publication statusPublished - 1 Jan 1990
Externally publishedYes

Fingerprint

alpha 1-Antitrypsin
Alleles
Isoelectric Focusing
Serum
Complementary DNA
Messenger RNA
Mutation
Emphysema
Amino Acid Substitution
Tamoxifen
Viperidae
DNA Sequence Analysis
Codon
Exons
Proteins
Fibroblasts
Genome
Phenotype

ASJC Scopus subject areas

  • Pulmonary and Respiratory Medicine

Cite this

Molecular analysis of the heterogeneity among the P-family of alpha-1-antitrypsin alleles. / Holmes, M. D.; Brantly, M. L.; Crystal, Ronald.

In: American Review of Respiratory Disease, Vol. 142, No. 5, 01.01.1990, p. 1185-1192.

Research output: Contribution to journalArticle

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abstract = "The rare P-family of α1-antitrypsin (α1AT) variants is defined by the position of migration of the α1AT protein on isoelectric focusing of serum (IEF) between the common M and S variants. To begin to examine the molecular heterogeneity among the P-type alleles, two unrelated subjects and their families identified by IEF to be carrying a P allele were analyzed. The first, P(lowell), is a deficiency allele associated with reduced serum α1AT levels, and the second, P(saint albans), is associated with normal serum levels. DNA sequence analysis of P(lowell), the more anodal of the two variants on IEF analysis, showed that it differed from the normal M1(Val213) allele by a single base and amino acid substitution Asp256 GAT → Val GTT. In contrast, P(saint albans), a slightly more cathodally positioned variant on IEF analysis, differed from the coding exons of the normal M1(Val213) allele by two mutations, Asp341 GAC → Asn AAC, and a silent substitution in the same codon as the P(lowell) variant, Asp256 GAT → Asp GAC. Evaluation of P(lowell) mRNA transcripts by Northern and cytoblot analyses demonstrated they were of normal size and amount, and P(lowell) mRNA transcripts could be translated normally in vitro. Retroviral insertion of the P(lowell) cDNA into the genome of 3T3 fibroblasts demonstrated that it directed the synthesis of α1AT, but at levels 24{\%} that of the P(saint albans) cDNA or the normal M1 (Val213) cDNA, with a pattern of biosynthesis consistent with the concept that the P(lowell) α1AT deficiency state results from intracellular degradation of the newly synthesized P(lowell) protein. In the context that the serum α1AT deficiency associated with other α1AT deficiency mutations resulting from intracellular degradation of α1AT can be overcome by administering estrogenlike drugs, administration of tamoxifen to a subject with the P(lowell)Z phenotype resulted in α1AT serum levels rising 48{\%} over a 5-month period, from below the threshold for protection from emphysema (11 μM) to above that threshold.",
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