PROTON-LINKED SUBUNIT KINETIC HETEROGENEITY FOR CARBON-MONOXIDE BINDING TO HEMOGLOBIN FROM CHELIDONICHTHYS KUMU

The pH dependence of CO binding kinetics to Chelidonichthys kumu hemog lobin (Hb) and human adult Hb has been investigated between pH 2.0 and 9.0 at 20 degrees C. For both Hbs, CO binding kinetics is characteriz ed by two proton linked transitions, with different pK(a) values for a lpha- and beta-chains in C. kumu Hb, leading to a relevant functional kinetic heterogeneity at most pH values, On the other hand, in human a dult Hb the CO binding does not display a functional heterogeneity, Lo wering the pH from 9 to 6 brings about a decrease of the CO binding ra te constants, to a different extent for human adult Hb and the two cha ins of C. kumu Hb, Further lowering the pH from 6 to 2 induces an enha ncement of CO binding rate constants, probably related to the protonat ion of proximal His(F8) N-epsilon atom and the cleavage (or severe wea kening) of the His(F8)-Fe bond, The presence of physiological concentr ations of ATP (approximate to 3 mM) affects the pH dependence of CO bi nding kinetics to C. kumu, Moreover, the effect of temperature (betwee n 8 degrees C and 38 degrees C) on CO binding kinetics has been invest igated in the absence of ATP at different pH values. These results all ow to interpret the functional kinetic heterogeneity of C. kumu Hb on the basis of different regulatory aspects in the alpha and beta subuni ts, as suggested by structural considerations.