标题：Characterization of Human p33/41 (Annexin IV), a Ca2+ Dependent Carbohydrate-Binding Protein with Monoclonal Anti-annexin IV Antibodies, AS11 and AS17
摘要：p33/41 (annexin IV) is a member of the family of Ca2+-dependent phospholipid binding proteins known as annexins. We previously described that bovine kidney p33/41 (annexin IV) has Ca2+-dependent carbohydrate binding acrivity. In this study, we purified human p33/41 (annexin IV) from the HT29, human colon adenocarcianoma cell line, as well as the bovine kidney annexin by affinity chromatography. Then, we prepared recombinant human p33/41 (annexin IV) expressed in Escherichia coli. The apparent size and the Ca2+-dependent carbohydrate binding properties of purified recombinant p33/41 (annexin IV) were indistinguishable from those of the bovine kidney protein. We also performed inhibition assays of carbohydrate binding and of phosphatidylserine/phosphatidylcholine liposome binding of recombinant p33/41 (annexin IV) with anti-p33/41 monoclonal antibodies (AS11 and AS17). We determined the epitopes recognized by the monoclonal antibodies by Western blot analysis using deleted-recombinant p33/41 (annexin IV). The monoclonal antibodies recognized domain 1 and/or 2 of p33/41 (annexin IV). The results of the inhibition assays and the determination of the epitope showed that a carbohydrate binding site is located at domains 3 and 4 of p33/41 (annexin IV) and on the cell surface.
