Gene action in the human haptoglobins. III. Isolation of the α chains as single gene products. Isolation molecular weight, and amino acid composition of α and β chains

Abstract

A modified method for the preparation of large quantities (1–10 g) of human haptoglobin (Hp) from the ascites fluid of a single patient has been developed. By selection of a heterozygous individual of genotype Hp1S/Hp2(F,S), and taking advantage of the different molecular sizes of the polypeptide gene products haptoglobin α1S and α2(F,S), it is possible to separate by Sephadex G-75 chromatography the reduced and alkylated polypeptide products of the single genes Hp1S and Hp2(F,S) for subsequent characterization and sequence analysis. The second polypeptide chain of haptoglobin, the β chain, is also obtained pure and in good yield by this procedure. The molecular weights of the β and α2 chains were determined after modification of the amino groups with succinic anhydride to reverse the extensive aggregation of reduced and alkylated β and α2 chains in neutral and alkaline solutions. Amino acid and carbohydrate compositions of the separated chains have been determined and it is shown that the empirical polypeptide chain constitution of haptoglobin 2–1 is (2 × β + α2 + α1).