ALPHA-SUBUNIT OXIDATION IN T-STATE CRYSTALS OF A SEBACYL CROSS-LINKEDHUMAN HEMOGLOBIN WITH UNUSUAL AUTOXIDATION PROPERTIES

In the crystal structure of human T-state hemoglobin with a sebacyl re sidue cross-linking the two beta-subunit Lys(82)'s (DecHb), the Fe ato ms of the alpha-subunit hemes are found to be oxidized with a water mo lecule bound. The three-dimensional structure and heme geometries were compared to those of deoxyhemoglobin and other partially and fully ox idized hemoglobins [R. Liddington, Z. Derewenda, E. Dodson, R. Hubbard , G. Dodson, J. Mel. Biol. 228 (1992) 551]. The heme geometries of the alpha-subunits are consistent with those observed in oxidized structu res, The proximal histidines of the cu-subunits move toward the heme p lane shifting the F-helix and FG-corner ina manner observed for partia lly oxidized human hemoglobin. This supports the hypothesis that these perturbations may precede the T-to R-state transition, Circular dichr oism studies comparing DecHb and natural human hemoglobin in the deoxy and CO ligated forms confirm that the conformations of the deoxy form s are identical, but the ligated forms have slight differences in the solution structures. DecHb is found to be more resistant to autoxidati on than natural hemoglobin. The time course of autoxidation of DecHb s hows that it is virtually absent for the first 1500 min followed by a rapid increase. Thus, the discovery of the oxidation of the cu-subunit s in the deoxy-crystals is quite unexpected. The data confirm that lig ation of the alpha-subunits precedes that of the beta-subunits. This m ay suggest a low ligand affinity of the alpha-diligated form of hemogl obin. (C) 1998 Elsevier Science B.V.