Processing of acidic proline-rich proprotein by human salivary gland convertase

Previously it was found that proproteins for basic and glycosylated salivary proline-rich proteins (PRP) were cleaved prior to secretion from cells by furin, a well-known convertase. In contrast proproteins for acidic PRPs are not cleaved by furin or other convertases. To investigate the convertase responsible for in vivo processing of acidic PRP proproteins, homogenates of human sublingual glands were fractionated by centrifugation at 10,000 x g and 100,000 x g and activity demonstrated in all fractions. The 100,000 x g pellet was fractionated into Golgi, smooth endoplasmic reticulum and microsomal fractions with the latter containing the enzyme. Subfractionation of the microsomes revealed that the activity was located in the membrane proteins. Since the microsomes contain components of the secretory pathway the enzyme in this fraction may be responsible for intracellular cleavage of the acidic PRP proprotein. The enzyme was active at alkaline pH. It was strongly inhibited by metal chelators indicating that it is a metalloprotease. It was not inhibited by an acid protease inhibitor, but partly inhibited by some serine protease inhibitors indicating that serine proteases may play a role in degradation. Co2+ and to some extent Zn2+ activated the enzyme, but it was strongly inhibited by Hg2+ and Cu2+ as well as the organomercurial p-chloromercuribenzenesulfonic acid. Thus it appears that the enzyme contains an important -SH group. These characteristics indicate that the convertase is related to a group of metal- and thiol-dependent proteases known as thimet oligopeptidases, but in contrast to the latter enzymes the sublingual convertase was not inhibited by angiotensin antagonists.