Purification, N-terminal sequencing, partial characterization, crystallization and preliminary crystallographic analysis of two glycosylated serine proteinases fromAgkistrodon acutusvenom

AaV-SP-I and AaV-SP-II, two glycosylated serine proteinases from Agkistrodon acutus venom with fibrinogenolysis and esterolysis activities, have been purified to homogeneity by three-step ion-exchange chromatography. Estimated by SDS-PAGE, the molecular weights of AaV-SP-I and AaV-SP-II are about 32 and 31 kDa under reducing conditions and 26 and 25 kDa under non-reducing conditions, respectively. The first 24 N-terminal amino-acid residues are the same in both sequences and display a high homology with those of several snake-venom serine proteinases. However, the proteins possess obviously distinct carbohydrate contents. Using the conventional hanging-drop vapour-diffusion method, single crystals of both enzymes were grown that were suitable for X-ray diffraction analysis. The crystals of AaV-SP-I and AaV-SP-II belong to space groups P2(1)2(1)2(1) and C2, respectively. In each case there is only one molecule in the asymmetric unit.