PACE4 is a member of the mammalian propeptidase family that has overlapping but not identical substrate specificity to PACE

Abstract

Proteins that transit the constitutive pathway of secretion frequently require proteolytic processing after a pair of basic amino acids to attain their full functional activity. A ubiquitously expressed calcium-dependent subtilisin-like serine protease, named PACE or furin, can cleave precursor polypeptides specifically at pairs of basic amino acids where an arginine residue is present in the P4 position. Another member of this protease family, PACE4, was cloned recently by a PCR-based strategy and was also shown to be ubiquitously expressed. We have expressed PACE4 by transient DNA transfection of COS-1 cells and have shown that the cDNA encodes a 120-kDa polypeptide that is present in cell extracts but not in conditioned medium of transfected cells. The substrate specificities of PACE and PACE4 for cleavage of pro-von Willebrand factor were studied in parallel using a transient DNA cotransfection system. Like PACE, PACE4 was able to process pro-vWF to its mature form, and efficient cleavage required both the P4 arginine and the P2 lysine. These data, taken together with previously published data showing that PACE4 cannot process pro-factor IX, demonstrate that PACE and PACE4 have overlapping but not identical substrate specificities. Further differences between PACE and PACE4 specificities were elucidated by monitoring inhibition of processing activity mediated by the serine protease inhibitor alpha 1-antitrypsin Pittsburgh mutant. Pro-vWF processing by PACE was inhibited by expression of the alpha 1-antitrypsin Pittsburgh mutant, whereas processing of pro-vWF by PACE4 was not affected.(ABSTRACT TRUNCATED AT 250 WORDS)

Cleavage of proalbumin peptides by furin reveals unexpected restrictions at the P2 and P′1 sites

“…The role that the furin homolog PACE4 may play in the liver is presently unclear but recent co-expression experiments have shown it to be proteolytically active and somewhat different to furin in its cleavage of mutated pro Von Willebrand factor (which is not produced in hepatocytes but endothelial cells). PACE4's lack of inhibition by g:antitrypsin Pittsburgh [20], however, makes it substantially different from furin and the in situ enzyme. The findings here, on the other hand, show that furin has a similar specificity to that expected of the in situ convertase.…”

PACE4: a subtilisin-like endoprotease with unique properties

“…However, no secretion of human PACE4 was detected from transfected COS-1 cells [32]. To investigate the synthesis and secretion of PACE4 more thoroughly, the hEK-293 line and additional stably transfected neuroendocrine and fibroblast cells were examined ( Figure 1).…”

“…Since PACE4 is encoded on the same chromosome as furin, and since some early reports suggested that PACE4 was not secreted and had a wide tissue distribution, the latter has been regarded as a soluble intracellular version of furin [32,61]. However, a further examination of PACE4 is important for several reasons.…”

“…For example, the inhibitory properties of the naturally occurring mutant α " -AT Pittsburgh (α " -AT-PITT), with a single Met Arg alteration, indicated that it was unable to abolish activity of purified furin [29]. When tested in cells, co-expression of α " -AT-PITT was unable to abolish furin-mediated processing of pro-β-nerve growth factor or PACE4-mediated cleavage of the von Willebrand precursor [32,60], although α " -AT-PITT inhibited furin-mediated processing of pro-von Willebrand factor [32]. Whereas furin cleaves the α-subunit of endopeptidase-24.18, PACE4 cannot [77].…”

Distinct mRNA expression of the highly homologous convertases PC5 and PACE4 in the rat brain and pituitary

“…PACE4 was also demonstrated to have intracellular processing activity by cleaving pro-von Willebrand factor (Creemers et al, 1993;Rehemtulla et al, 1993). The catalytic domain of PACE4 exhibits 70% identity to that of PC5.…”