Structural and Functional Characterization of Haemocyanin from the Anemone Hermit Crab Dardanus calidus

Oxygen-binding to haemocyanin (Hc) is generally an exothermic process, with overall enthalphy of oxygenation varying from species to species. A number of crustacean Hcs showed a null or reduced enthalphy of oxygenation, among others, the anomuran Pagurus bernhardus and Paralithodes camtscaticae possess a completely temperature-independent oxygen-binding in a wide range of temperature and pH. Functional analysis performed on purified native, hexameric and dodeca-meric Hc forms of the anemone hermit crab Dardanus calidus allowed to calculate the enthalphy of oxygenation values that resulted equal to 236.2, 233.8 and 226.8kJ/ mol, respectively. Thus, the temperature sensitivity of oxygen binding of D. calidus Hc is in contrast with the temperature independence reported for P. bernhardus and P. camtscaticae, suggesting a high Hc functional heterogeneity within Anomura. Functional characterization also evidenced a strong oxygen affinity modulation by protons ("logP50/"pH=20.97) and lactate ["logP50/"log(lactate) =20.38], and a significant decrease in cooperativity by physiological concentration of lactate (n50 from 2.8 to 1.7 at pH 7.5)