Although the molecular defect in
sickle hemoglobin that produces
sickle cell disease has been known for decades, there is still no effective
drug treatment that acts on
hemoglobin itself. In this work, a series of diversely substituted
isothiocyanates (R-NCS) were examined for their regioselective reaction with
hemoglobin in an attempt to alter the solubility properties of
sickle hemoglobin. Electrospray mass spectrometry, molecular modeling, X-ray crystallography, and conventional
protein chemistry were used to study this regioselectivity and the resulting increase in solubility of the modified
hemoglobin. Depending on the attached R-group, the
isothiocyanates were found to react either with the Cysbeta93 or the N-terminal
amine of the alpha-chain. One of the most effective compounds in the series, 2-(N,N-dimethylamino)ethyl
isothiocyanate, selectively reacts with the
thiol of Cysbeta93 which, in conjunction with the cationic group, was seen to perturb the local
hemoglobin structure. This modified HbS shows an approximately 30% increase in solubility for the fully deoxygenated state, along with a significant increase in
oxygen affinity. This compound and a related analogue appear to readily traverse the erythrocyte membrane. A discussion of the relation of these structural changes to inhibition of gelation is presented. The dual activities of increasing HbS
oxygen affinity and directly inhibiting
deoxy HbS polymerization, in conjunction with facile membrane traversal, suggest that these cationic
isothiocyanates show substantial promise as lead compounds for development of therapeutic agents for
sickle cell disease.