beta 93 modified hemoglobin: Kinetic and conformational consequences

The reactive sulfhydryl on Cys beta 93 in human adult hemoglobin (HbA) has been the focus of much attention. It has purported functional roles such as a transporter of nitric oxide and a detoxifier of super oxide. In addition , it has a proposed role in the allosteric mechanism. The present study add resses the functional and conformational consequences of modifying the beta 93 sulfhydryl using either maleimide or disulfide-based reactions. The gem inate and bimolecular recombination of CO derivatives of several different beta 93-modified Hbs in both solution and sol-gel matrixes provide a window into functional modifications associated with both the R and T states of t hese proteins. Nanosecond time-resolved visible resonance Raman spectroscop y is used to probe conformational consequences associated with the proximal heme environment, The results show functional and conformational consequen ces that depend on the specific chemistry used to modify beta 93. Maleimide -based modification show the most significant alterations of R-state proper ties including a consistent pattern of a reduced geminate yield and a loss of the favorable heme-proximal histidine interaction normally seen for liga nded R-state HbA. A mechanism based on a displacement of the side chain of Tyr beta 145 is explored as a basis for this effect as well as other situat ions where there is loss of the quaternary enhancement effect. The quaterna ry enhancement effect refers to the enhancement of ligand binding propertie s of the alpha beta dimers when they are associated into the R-state tetram er.