PC4 or PCSK4 belongs to the 9-member superfamily of mammalian subtilases collectively called
Proprotein Convertases or Proprotein Convertase Subtilisin/Kexins that convert inactive precursor proteins into
their active mature forms by endoproteolytic cleavage. PC4-activity plays a crucial role in mammalian
fertilization via activation of sperm surface proteins. PC4 knockout mice exhibit severely impaired male fertility
due to premature sperm acrosome reaction. Regulation of sperm-PC4 activity during its storage and transport
through epididymis is an important determinant for ultimate egg-binding and fertilizing capacities of sperms.
Herein we show that epididymal serpin CRES (cystatin related epididymal spermatogenic) recombinant protein
inhibits PC4 activity in vitro in a differential manner when measured against the fluorogenic substrate Boc-
RVRR-MCA depending on its oligomeric state. Thus while CRES-dimer exhibits Ki ~8 μM, the corresponding
monomer showed Ki > 100 μM. Both forms also blocked PC4-mediated processing of human proIGF-2 in
human placenta tropoblast cell line with dimer being more efficient. Using specific inhibitors and substrates, we
also demonstrated the presence of PC4-like activity and CRES protein in varying levels in the fluids of various
epididymal compartments. Our observations suggest a potential function of CRES as a regulator of PC4 in
sperm-egg interaction and fertilization.
Keywords: Proprotein convertase 4, proprotein convertase subtilisin kexin 4, serpin, epididymal fluid, cystatin
related epididymal spermatogenic, inhibitor, protein aggregation, proprotein processing, yeast kexin, bacterial subtilisin, proneuropeptides, prohormones, maturation, autocatalysis, fertilization.
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