Further Improvement in Preparation and Some Properties of Native α and β Chains from Canine Hemoglobin Treated with p-Chloromercuribenzoate

• Published in: Journal of biochemistry (Tokyo) Vol. 96; no. 4; pp. 985 - 991
• Main Authors: TOMITA, Susumu, SAKATA, Susumu, ENOKI, Yasunori, KOHZUKI, Hisaharu
• Format: Journal Article
• Language: English
• Published: Oxford Oxford University Press 01.10.1984
• Subjects: Amino Acids - analysis; Analytical, structural and metabolic biochemistry; Animals; Biological and medical sciences; Chloromercuribenzoates; Dogs; Electrophoresis, Starch Gel; Fundamental and applied biological sciences. Psychology; Heme - analysis; Hemoglobins - isolation & purification; Hemoproteins; Humans; Macromolecular Substances; Metalloproteins; p-Chloromercuribenzoic Acid; Protein Conformation; Proteins; Species Specificity; Spectrophotometry; Sulfhydryl Compounds - analysis; Fissipedia; Carnivora; Hemoprotein; Oxygen; Beta-Peptide chain; Method; Absorption spectrometry; Improvement; Vertebrata; Mammalia; Animal; Hemoglobin; Affinity; Alpha-Peptide chain; Isolation; Dog
• ISSN: 0021-924X; 1756-2651
• DOI: 10.1093/oxfordjournals.jbchem.a134957

The α and β chains were prepared from canine hemoglobin by a modification of the method of Bucci and Fronticelli ((1965) J. Biol. Chem. 240, PC551–552) which involved treatment of hemoglobin with an excess of p-chloromercuribenzoate (pCMB), separation of the mercurated chains by chromatography on a DE-32 column65 with a salt gradient at pH 8.6, and regeneration of sulfhydryl groups of the chains with 2-mercaptoethanol. The SH titer was two per heme for both the regenerated α and β chains. The titer decreased to four per tetramer of hemoglobin after equimolar recombination of both chains. Measurements of the absorption spectrum and oxygen binding showed that the chains were in a native state.