Proteinase Activity of Human Renin Preparations: The International Reference Preparation (Renin Standard 68/356)

Abstract

1. Several commonly used preparations of human renin, including the International Reference Preparation (Renin Standard 68/356), were examined for the presence of contaminating proteinase activity by using a 14C-glycinated bovine haemoglobin substrate assay. 2. All of the human renin preparations tested cleaved haemoglobin even in the presence o di-isopropylfluorophosphate and ethylenediaminetetra-acetate (EDTA). For a given amount of renin activity, varying amounts of proteinase activity were seen. The pH optimum also varied between preparations. 3. Small peptide inhibitors of human renin were not able to inhibit the proteinase activity. Furthermore a diazoacyl reagent and pepstatin, both potent inhibitors of aspartic acid-active site proteinases, were only partially inhibitory. 4. These and other observations suggest that the proteinase activity of the human renin preparations is not due to renin itself, but to contaminating proteinases of different types. Since these enzymes may produce angiotensin I or peptides which may interfere in renin assays, crude preparations of renin which contain proteinase activity, including the International Reference Preparation, should be used with caution or replaced by proteinase-free human renin which can be easily prepared by use of suitable affinity chromatography.

Pepstatin Inhibition of Proteinase-Free Human Renin

“…We have earlier referred to the probability that the proteinase(s) contami nating renin preparations ('pseudorenin') were likely to be cathepsin-D or some similar enzymes [3], Clearly, however, this is not the case. When pepstatin inhibition of cathepsin-D was studied, tight-binding kinetics were observed [2], Furthermore, the proteinases contaminating crude renin preparations are not completely inhibited by a diazoacyl re agent which does inhibit cathepsin-D [5]. The proteinase contaminants of crude renin are clearly multiple, and the earlier results suggest that cathepsin-D or other acid pro teinases are not the only contaminants.…”