ISOMERIZATION REACTIONS OF CHARCOAL-DEFATTED BOVINE PLASMA ALBUMIN . N-F TRANSITION AND ACID EXPANSION

In view of the possibility that bound lipophilic impurities might make an important contribution to the observed microheterogeneity of bovine plasma albumin (BPA) samples, the character of the population distributions of albumin defatted by the conventional acid procedure and by the newer charcoal procedure has been determined by means of solubility-pH profiles and comparison was made with the nondefatted albumin used as precursor to these preparations. The results suggest that acid defatting leads to an artificial broadening of the population due to two factors, namely (1) partial but incomplete defatting and (2) some unknown irreversible alteration of some of the protein. The charcoal-defatted protein has a much narrower population distribution than either nondefatted or acid-defatted BPA. Improved procedures for determining solubility-pH profiles and for subfractionation of the populations were developed. The most striking property of the charcoal-defatted protein is its lability toward some reaction which results in broadening of the population. This process is most rapid above pH 7 but is appreciable even at the isoionic pH and at more acid pH in deionized solution, but is retarded by adding salt at concentrations as low as 0.03 m. Owing to the more homogeneous population, the acid transitions of charcoal-defatted protein are sharper and better resolved than in previous preparations. The N-F transition and acid expansion can be clearly resolved by optical rotation measurements at the trough of the first Cotton effect, 233 mμ, and by pH difference spectra. Careful measurements have been made of the optical rotatory dispersion properties of charcoal- defatted bovine plasma albumin both at neutral pH and through the range of the acid transitions. Hydrogen ion titration results are presented for the acid range which agree in general with previously published results but the number of titratable carboxyl groups (102) is in better agreement with amino acid composition than the number found earlier (108) for acid-defatted protein, suggesting the possibility that some deamidation may accompany the low pH treatment. © 1968, American Chemical Society. All rights reserved.