SUBUNIT STRUCTURE AND OXYGEN BINDING BY HEMOCYANIN OF THE ISOPOD LIGIA-EXOTICA

Isopods are small crustacean arthropods that are generally found in damp or humid areas. The “rock louse”, “sow bug”, “pill bug”, and “wood louse” are members of this group. While the hemocyanins of crabs and shrimps have been the object of numerous investigations, very little work has been done on isopod hemocyanins. This is undoubtedly due, in part, to the small size of the isopods which makes it difficult to obtain hemocyanin samples in appreciable quantity. We have examined in detail the hemocyanin of the marine isopod Ligia exotica. The molecular architecture of the protein resembles that of decapod crustaceans in that dodecamers and hexamers are found in the hemolymph. Only the hexameric form persists after removal of divalent cations by treatment with EDTA at pH 9.0. Monomeric forms appear at elevated pH and molecular sieve chromatography can be used to separate hexamers and monomers. At a given pH the isolated fractions are stable populations of either monomers or hexamers, implying the existence of multiple pK values for hexamer-monomer equilibria. At pH 9.5 the isolated monomeric subunits have lower oxygen affinities than the hexamers, indicating that in this hemocyanin system the aggregation of subunits introduces constraints which lead to increased oxygen affinity. The monomers which are isolated by molecular sieve chromatography give single bands on regular and NaDodSO4 gels and are thus considered to be homogeneous with respect to charge and molecular weight. They reaggregate at neutral pH and the reassembled hexamers show cooperative interactions in oxygen binding. The high degree of cooperativity and positive Bohr effect exhibited by hexameric aggregates made up of apparently identical subunits make Ligia hemocyanin a good model system for investigation of homo- and heterotropic interactions in a high molecular weight system. © 1979, American Chemical Society. All rights reserved.