Hemoglobin subunit alpha
oxygen transport
Expression, purification, and characterization of human hemoglobins Gower-1 (zeta(2)epsilon(2)), Gower-2 (alpha(2)epsilon(2)), and Portland-2 (zeta(2)beta(2)) assembled in complex transgenic-knockout mice.
Embryonic zeta- and epsilon-globin subunits assemble with each other and with adult alpha- and beta-globin subunits into hemoglobin heterotetramers in both primitive and definitive erythrocytes. The properties of these hemoglobins-Hbs Gower-1 (zeta(2)epsilon(2)), Gower-2 (alpha(2)epsilon(2)), and Portland-2 (zeta(2)beta(2))-have been incompletely described as they are difficult to obtain in quantity from either primary human tissue or conventional expression systems. The generation of complex transgenic-knockout mice that express these hemoglobins at levels between 24% and 70% is described, as are efficient methods for their purification from mouse hemolysates. Key physiological characteristics-including P(50), Hill coefficient, Bohr effect, and affinity for 2,3-BPG-were established for each of the 3 human hemoglobins. The stability of each hemoglobin in the face of mechanical, thermal, and chemical stresses was also determined. Analyses indicate that the zeta-for-alpha exchange distinguishing Hb Portland-2 and Hb A alters hemoglobin O(2)-transport capacity by increasing its P(50) and decreasing its Bohr effect. By comparison, the epsilon-for-beta exchange distinguishing Hb Gower-2 and Hb A has little impact on these same functional parameters. Hb Gower-1, assembled entirely from embryonic subunits, displays an elevated P(50) level, a reduced Bohr effect, and increased 2,3-BPG binding compared to Hb A. The data support the hypothesis that Hb Gower-2, assembled from reactivated epsilon globin in individuals with defined hemoglobinopathies and thalassemias, would serve as a physiologically acceptable substitute for deficient or dysfunctional Hb A. In addition, the unexpected properties of Hb Gower-1 call into question a common hypothesis for its primary role in embryonic development.