THERMAL-STABILITY OF HEMOGLOBIN CROSS-LINKED IN THE T-STATE BY BIS(3,5-DIBROMOSALICYL) FUMARATE

Bis(3,5-dibromosalicyl) fumarate was used to crosslink oxyhemoglobin between Lys 82β1 and Lys 82β2 (Walder, J. A., et al. (1979) Biochemistry 18, 4265) and deoxyhemoglobin between Lys 99α1 and Lys 99α2 (Chatterjee R.Y., et al.(1986) J. Biol. Chem. 261, 9929). Thermal denaturations demonstrated that α crosslinked hemoglobin (α99XLHb A) has the same stability as the β crosslinked one (β82XLHb A). Both α and β crosslinked methemoglobins have a denaturation temperature in 0.9 M guanidine of 57°C compared to 41°C of Hb A. The second product from the T-state crosslinking reaction was found to be crosslinked between the β chains by chain separation and amino acid analysis. The possible positions for this crosslink are limited to the bisphosphoglycerate binding site in the three-dimensional structure. Its stability is comparable to that of the α99XLHb A or β82XLHb A. These modified hemoglobins are potential blood substitutes. © 1991.