Two-step affinity-chromatographic purification of cathepsin D from pig myometrium with high yield

Abstract

Cathepsin D was purified by two-step affinity chromatography on concanavalin A- and pepstatin-Sepharose. The main purification was achieved by washing the enzyme bound to the pepstatin-Sepharose column with buffered 6 M-urea. This step separated cathepsin D from all low- and high-molecular-weight impurities. Although the 1700-fold purified acid proteinase was homogeneous on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, it still showed microheterogeneity.

Effective blocking of HIV‐1 proteinase activity by characteristic inhibitors of aspartic proteinases

“…5.5 has been observed previously with typical aspartic proteinases (e.g. [13]) and indeed has been used to effect in developing affinity chromatographic procedures for the convenient purification of the enzymes [19]. H-261, however, remained an adequate inhibitor (table 1) with an ICSO value of 25 nM estimated at pH 7.0.…”

Human cathepsin D.

“…The method of cathepsin D isolation from the human liver [29][30][31][32][33], spleen [18], brain [37], uterus [34], placenta [35], gastric mucosa [36], thyroid [38] and leucocytes [39] has been described. The human liver cathepsin D preparation is produced by Calbiochem and Sigma.…”

Characterization of pregnancy-associated glycoproteins extracted from zebu (Bos indicus) placentas removed at different gestational periods

“…Pepstatin A is an extremely powerful inhibitor of aspartic proteinases [14]. It binds reversibly to the active center of pepsin, renin and cathepsin D, being widely used as a ligand in affinity chromatography protocols for the isolation of these enzymes [1,5]. In the present work, PAG molecules were successfully identified after pepstatin affinity chromatography of cotyledonary extracts from placentas collected in both the first and second trimesters of gestation and extracted in both acid and alkaline conditions.…”