Incorporation of low molecular weight molecules into α2-Macroglobulin by nucleophilic exchange

α2-Macroglobulin (α2M) is a proteinase inhibitor that functions by a trapping mechanism which has been exploited such that the receptor-recognized, activated form (α2M∗) can be employed to target antigens to antigen-presenting cells. Another potential use of α2M∗ is as a drug delivery system. In this study we demonstrate that guanosine triphosphate, labeled with Texas red (GTP-TR) formed complexes with α2M∗ following activation by proteolytic or non-proteolytic reactions. Optimal incorporation occurred with 20 μM GTP-TR, pH 8.0 for 5 h at 50 °C. NaCl concentration (100 or 200 mM) had little effect on incorporation at this pH or temperature, but was significant at sub-optimum temperature and pH values. Maximum incorporation was 1.2 mol GTP-TR/mol α2M∗. PAGE showed that 70-90% of the GTP-TR is bound in a SDS/2-mercaptoethanol resistant manner. Guanosine, adenosine, and imidazole competed with GTP-TR to form complexes with α2M∗.