Functional effects of replacing human alpha- and beta-globins with their embryonic globin homologues in defined haemoglobin heterotetramers

Embryonic- and adult-stage globin subunits assemble into haemoglobin (Hb) h eterotetramers that are expressed at low levels throughout human intrauteri ne development. These haemoglobins differ from adult Hb A (alpha(2)beta(2)) by the substitution of embryonic zeta for adult alpha globin (Hb zeta(2)be ta(2)), or embryonic epsilon for adult beta globin (Hb alpha(2)epsilon(2)). Several key physiological properties of these 'semiembryonic' haemoglobins remain undefined, as ethical and methodological considerations have limite d their availability from both human sources and conventional expression sy stems. The current study attempts to estimate how the physiological propert ies of semiembryonic and adult haemoglobins may differ, by determining whet her the O-2-binding characteristics of hybrid human/mouse haemoglobins chan ge when human alpha- or beta-globin subunits are replaced by human embryoni c zeta- or epsilon-globin subunits respectively. Each of the four human glo bins is expressed in transgenic mice that are nullizygous for either the en dogenous mouse alpha- or beta-globin genes, resulting in the high-level exp ression of haemoglobins that can be studied either in situ in intact erythr ocytes or in vitro. We showed that the exchange of human zeta-globin for hu man alpha-globin chains increased haemoglobin O-2 affinity, both in the pre sence and in the absence of 2,3-bisphosphoglycerate (2,3-BPG), and reduced the pH-dependent shift in its oxygen equilibrium curve (Bohr effect). By co mparison, hybrid haemoglobins containing either human epsilon-globin or hum an beta-globin exhibited nearly identical O-2-binding properties, both in s itu and in vitro, regardless of 2,3-BPG levels or ambient pH. Neither the z eta-for-alpha nor the epsilon-for-beta substitutions substantially altered binding affinity for 2,3-BPG or cooperativity between globin subunits. Thes e studies suggest that semiembryonic haemoglobins that assemble entirely fr om human subunits may exhibit properties that are similar to those of human Hb A.