title: Heterogeneous binding of oxygen and carbon monoxide to dissociated molluscan hemocyanin.
abstract: Functional heterogeneity in O2 or CO binding of sites of dissociated molluscan hemocyanin polypeptide chains (Helix pomatia and Octopus vulgaris) has been estimated by an analysis of accurate noncooperative binding curves. Three types of experiments were performed: pure O2 or CO binding, competitive displacement of one ligand by the other, and simultaneous removal of both gases from protein partially saturated with O2 and CO. The data were analyzed in terms of a model which has two fractions of sites with different properties for O2 and CO. The relative proportion of the different binding sites and their affinity constant values were found by the combined use of the three different procedures. All species show a marked functional heterogeneity of sites for O2 binding, while for CO binding it has been observed only in the case of H. pomatia beta-hemocyanin. Moreover, in all three molluscan hemocyanins examined, the two classes of O2-binding sites, although present in different proportions within the polypeptide chains, display similar affinity constant values. The data reported show a good consistency with results obtained using digested and isolated domains, providing confidence in the analytical procedure used. From comparison of the O2/CO affinity ratios (KO2, KCO) of each class it may be suggested that the difference in O2 affinity of two kinds of binding sites is related to a different local structure of the active sites. The results, moreover, unequivocally confirm that binding and displacement of two gaseous ligands to hemocyanin occur by a simple competitive mechanism, although the binding site is structurally complex and the two ligands are bound with different geometries.
authors.affiliation: Institute of Chemistry, Faculty of Medicine and CNR Center of Molecular Biology, University of Rome 'La Sapienza', 00185 Rome, Italy.
authors.forename: L
authors.initials: L
authors.lastname: Zolla
authors.forename: M
authors.initials: M
authors.lastname: Brunori
authors.forename: B
authors.initials: B
authors.lastname: Richey
authors.forename: S J
authors.initials: SJ
authors.lastname: Gill