Human α1-Antitrypsin Characterization and N- and C-Terminal Sequences1

Abstract

Two forms of alpha-1-antitrypsin (α₁-AT), α₁-AT(T), and α¹-AT(lT), were separately obtained from human sera which had been pooled and stored for 3 to 6 months at −20°C by a combination of ion-exchange chromatography, gel-filtration, and affinity chromatography. The N-terminal amino acid sequences of α¹-AT(I) and α¹-AT(II) were tentatively determined as Glu-Asp-Pro-Gin-Gly-Asp-Ala-Ala-Gln-Glu-Thr-Asp-Thr-Ser-His-(His)- and Glu-Thr-Asp-Thr-Ser-His-His-Asp-GIn-Asp-Ser-Pro-Thr-Phe-Asn-Lys-Leu-Thr-Pro-Asn-Leu-Ala-Glu-Phe-, respectively. The amino acid sequence of α¹AT(lI) corresponds to the sequence beginning at the tenth glutamic acid residue from the N-terminus of α¹-AT(l). Thus, α¹-AT(ll) may arise by cleavage of the Gln-Glu bond in α—AT(I) by an enzyme either originally present in the serum or produced by contaminating microorganisms. Based on a combination of the N-terminal sequences of α¹AT(I) and α¹-AT(ll), the sequence of the N-terminal 33 residues of α¹-AT is tentatively proposed. α¹-AT isolated from fresh human plasma had the same N-terminal amino acid sequence as αAT(I). The C-terminal amino acid sequences of the three α¹AT‘s were identified as -Gln-Lys by the carboxypeptidase digestion method. Amino acid analysis of performic acid-oxidized α¹-AT showed that this protein contained one mole of cysteine residue per mole of protein, which was linked to a free cysteine through a disulfide bond. Most of the α¹-AT preparations inhibited bovine trypsin at a molar ratio of 1: 1. However, certain preparations bound more than one mole of trypsin per mole of inhibitor.