KEYHOLE LIMPET HEMOCYANIN - STRUCTURAL AND FUNCTIONAL-CHARACTERIZATION OF 2 DIFFERENT SUBUNITS AND MULTIMERS

Keyhole limper hemocyanin (KLH), the large respiratory glycoprotein fr om the primitive gastropod mollusc, Megathura crenulata, is a potent i mmunogen used classically as a carrier protein for haptens and more re cently in human vaccines and for immunotherapy of bladder cancer. Two KLH isoforms were identified and isolated by high-performance anion ex change chromatography. Subsequent analyses disclosed that these isofor ms-designated KLH-A and KLH-B-were composed oi distinct subunits that differed in primary structure, molecular weight (KLH-A was 449,000 and KLH-B was 392,000:), polymerization/reassociation characteristics, an d O-2-binding constants (KLH-A had a P-50 Of 7.32 and KLH-B had a P-50 Of 2.46). Both subunits appear to be composed of eight oxygen binding domains, and reassociate in solution only with like subunits. These r esults support the concept that structural and functional heterogeneit y is a common feature of molluscan hemocyanins, and provide a rational basis for studying and optimizing the immunostimulatory properties of KLH.