Regulation of human MBL-MASPs by C1-inhibitor and α2-macroglobulin

Abstract

IL-12 is secreted by macrophages and antigen presenting cells, and is a key cytokine at the interface between innate and acquired immunity, initiating cell-mediated (Thl -type) immune responses. We have previously shown using an ELISA method, that recombinant IL-12, both human and murine, binds specifically and with high affinity to heparin and heparan sulphate glycosaminoglycans (Hasan et al. J. Immunol. 1999Immunol. 162: 1064. IL-12 is a heterodimer, comprising 40 and 35kD subunits. We found that the p40 subunit in the absence of p35 also binds strongly to heparin. In order to characterise the interaction between IL-12 and glycosaminoglycans further, we have now found that N-desulphation, selective 6-0-desulphation, and selective 2-0-desulphation of heparin all result in preparations which show considerably reduced, but not abolished, binding. We therefore conclude that each of these three classes of sulphate plays some partial role in binding. We have also examined the ability of heparin and heparan sulphate to protect rIL-12 from proteolytic digestion in vitro. Both subunits are protected by heparin from degradation by the lysine-specific enzyme, Lys C, but the effect on the p40 subunit is particularly marked. The physiological importance of this protease protection now needs to be established. We are continuing to investigate the structural basis and functional consequences of the binding of IL-12 to heparin and heparan sulphate. 221 9 Regulation of human MBL-MASPs by CI-inhibitor and a2macroglobulin M.