CALORIMETRIC EVIDENCE FOR ALLOSTERIC SUBUNIT INTERACTIONS ASSOCIATED WITH INHIBITOR BINDING TO BAND-3 TRANSPORTER

A calorimetric endotherm occurring at 68-degrees-C (the C-transition) has been assigned previously to the integral domain of band 3 and was shown to be shifted to 78-degrees-C after covalent binding of 4,4'-dii sothiocyanostilbene-2,2'-disulfonate (DIDS). In this study, we correla te the fractional appearance of the shifted C-transition with the frac tion of DIDS bound to the band 3 monomer popuIation. Our results show a distinctly nonlinear correlation plot with the appearance of the shi fted C-transition lagging behind DIDS labeling of the band 3 monomer p opulation. The lag suggests that both monomers of a band 3 dimer must be labeled by DIDS in order for the shifted C-transition to appear at 78-degrees-C, implying that the thermal unfolding of the integral doma in of band 3 is modulated by allosteric interactions between subunits. This is the first in situ structural evidence supporting ligand-media ted subunit interactions within a ''carrier''-type transporter protein oligomer.