Spectrophotometric Titrations of Human Serum Albumin and Reduced Carboxymethylated Albumin

Abstract

The tyrosyl groups of reduced and carboxymethylated human serum albumin, in which all disulfide bonds are broken, ionize at lower pH than those of native albumin, in spite of the greater negative charge on the protein produced by the S-carboxy-methyl groups. The heat and entropy of ionization of these groups in the reduced albumin are also "normal," in contrast to native albumin. Spectro-photometric titrations at 244 and 295 mµ give equivalent results. The more open structure of the reduced albumin, as compared with the native protein, presumably allows the tyrosine groups to come freely into contact with solvent. In acid solutions the heights of the two peaks at 236 and 287 mµ in the difference spectrum (albumin at pH 7 against albumin at a lower pH) are strictly proportional over the whole course of the titration. Both peaks apparently arise from changes in the environment of tyrosyl residues, rather than from more general conformational changes.