Abstract
By tandem-crossed immunoelectrophoresis and ELISA experiments an immunological relationship was observed between a-macroglobulin (alpha M) and hemocyanin (Hc) of the terrestrial snail Helix pomatia. Both glycoproteins occur in the hemolymph: alpha M (minor component) as a specific proteinase inhibitor, Hc (consisting of three components: alpha(D)-HpH, alpha(N)-HpH and beta-HpH) as oxygen transport protein. The cross-reaction was found to be correlated with glycosylation. (i) With beta-HpH, which is richer in carbohydrates than alpha(D)-HpH and alpha(N)-HpH, mainly due to a higher 3-O-methyl-D-galactose content, the cross-reaction with Hp alpha M was highest. (ii) From the 8 functional units, designated a-h, isolated from beta-HpH, two that lack carbohydrates (c and f) were not recognized by antibodies against Hp alpha M, while the six glycosylated ones were strongly cross-reacting. The nearly complete loss of the cross-reactivity upon deglycosylation of functional units d and g and the inhibition in competitive ELISA experiments by glycopeptides isolated from both beta-HpH and HpaM are further evidence that glycans are involved in the immunological relationship between HpH and Hp alpha M. Carbohydrate analyses indicated that the glycan structures present on HpaM are very similar (or identical) to those found on HpH, suggesting that glycans are common epitopes on both proteins. Especially D-xylose and 3-O-methyl-D-galactose seem to be responsible for the cross-reactivity since the m-macroglobulin and hemocyanin of the cephalopod Sepia officinalis, which lack these two monosaccharicles in their glycan structures, do not immunologically cross-react. (C) 2009 Elsevier Masson SAS. All rights reserved.