Scientific Publications by FDA Staff

Biochem J 2017 Dec 11;474(24):4171-92

Engineering Oxidative Stability in Human Hemoglobin based on the Hb Providence (betaK82D) mutation and Genetic Crosslinking.

Strader MB, Bangle R, Parker Siburt CJ, Varnado CL, Soman J, Benitez Cardenas AS, Samuel PS, Singleton EW, Crumbliss AL, Olson JS, Alayash AI

Previous work suggested that hemoglobin (Hb) tetramer formation slows autoxidation and hemin loss and that the naturally occurring mutant, Hb Providence (ßK82D) is much more resistant to degradation by H2O2 We have examined systematically the effects of genetic crosslinking of Hb tetramers with and without the Hb Providence mutation on autoxidation, hemin loss, and reactions with H2O2, using native HbA and various wild-type recombinant Hbs as controls. Genetically crosslinked Hb Presbyterian (ßN108K) was also examined as an example of a low oxygen affinity tetramer. Our conclusions are: (a) at low concentrations, all the crosslinked tetramers show smaller rates of autoxidation and hemin loss than HbA, which can dissociate into much less stable dimers and (b) the Hb Providence ßK82D mutation confers more resistance to degradation by H2O2, by markedly inhibiting oxidation of the ß93 cysteine side chain, particularly in crosslinked tetramers and even in the presence of the destabilizing Hb Presbyterian mutation. These results show that crosslinking and the ßK82D mutation do enhance the resistance of Hb to oxidative degradation, a critical element in the design of a safe and effective oxygen therapeutic.