Assembly and calcium-induced cooperativity of Limulus IV hemocyanin: a model system for analysis of structure-function relationships in the absence of subunit heterogeneity.
Hemocyanins, the high molecular weight copper proteins which serve as oxygen carriers in many arthropods and molluscs, are representative of multisubunit complexes which are capable of reversible dissociation and assembly. Although reversible, in many hemocyanins these processes are not in true thermodynamic equilibria, and it has been suggested that there is "microheterogeneity" among the molecules in solution. An alternative explanation is that their complex behavior is due to the existence of quaternary interactions between structurally distinct types of subunits within the native molecule which have varying pH and ionic strength sensitivity. Limulus IV hemocyanin was used as a model system to examine structure-function relationships in the absence of subunit heterogeneity. Purified subunit IV of Limulus polyphemus hemocyanin is homogeneous by a number of electrophoretic and immunological criteria and is capable of undergoing pH-dependent self-assembly into hexamers. The monomer-hexamer transition was found to be an equilibrium whose rate is dependent on the presence or absence of calcium ions. The observation that the assembly of this homopolymer behaves as a true equilibrium suggests that the nonequilibrium dissociation profiles observed for native Limulus hemocyanin are related to the extensive subunit heterogeneity of the native protein. In calcium-containing buffers, the monomer-hexamer transitions of Limulus IV hemocyanin can be described by a cooperative mechanism with approximately six protons per hexamer lost on assembly from acid pH and three protons gained on assembly from alkaline pH. Increased ionic strength or increased temperature favors dissociation. Like the native molecule, Limulus IV hemocyanin behaves as an allosteric protein.(ABSTRACT TRUNCATED AT 250 WORDS)
D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D006433:Hemocyanins / Q000378:metabolism*; D006737:Horseshoe Crabs / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D046911:Macromolecular Substances; D008433:Mathematics; D008954:Models, Biological; D013329:Structure-Activity Relationship
D046911:Macromolecular Substances; D006433:Hemocyanins; D002118:Calcium