Analysis of bicarbonate binding to crocodilian hemoglobin.
Crocodilian hemoglobin has a high intrinsic oxygen affinity but does not react with those organic phosphate esters that normally control the oxygen affinity of blood in higher vertebrates. Instead, its oxygen affinity is greatly lowered by CO2. The present study was undertaken to determine the nature of the CO2 binding to the hemoglobin of a crocodilian species, the Caiman, both qualitatively and quantitatively. The following parameters were measured: (a) carbamino compounds of deoxy- and oxyhemoglobin, (b) the effect of CO2 (at constant pH) on the oxygen affinity of Caiman hemoglobin, (c) total CO2 concentration of hemoglobin solutions at different pH and pCO2 values, and (d) the effect of CO2 on CD spectra of Caiman aquomethemoglobin. An analysis of the results of these measurements revealed that CO2 binding in the form of carbamate was not oxygen-linked and cannot, therefore, mediate the CO2 effect on the oxygen affinity. It was found, however, that 2 mol of bicarbonate can be bound/hemoglobin tetramer and that the association constant of the bicarbonate anion greatly depends upon the state of ligation. At pH 7.02 and 25 degrees C, a numerical value of 2.0 X 10(3) M-1 was obtained for deoxyhemoglobin, while for oxyhemoglobin no significant bicarbonate binding could be observed. At more alkaline pH (pH greater than or equal to 7.5), the association constant for deoxyhemoglobin decreases. Circular dichroism of Caiman aquomethemoglobin decreased considerably in the 287-nm region upon addition of CO2 at constant pH, an effect very similar to the one caused by inositol hexaphosphate in human aquomethemoglobin.
D000489:Alligators and Crocodiles / Q000097:blood*; D000818:Animals; D001639:Bicarbonates / Q000097:blood*; D002942:Circular Dichroism; D006454:Hemoglobins / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D010108:Oxyhemoglobins / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D012104:Reptiles / Q000097:blood*
D001639:Bicarbonates; D006454:Hemoglobins; D010108:Oxyhemoglobins