Implantation serine proteinase 2 is a monomeric enzyme with mixed serine proteolytic activity and can silence signalling via proteinase activated receptors.
Implantation serine proteinase 2 (ISP2), a S1 family serine proteinase, is known for its role in the critical processes of embryo hatching and implantation in the mouse uterus. Native implantation serine proteinases (ISPs) are co-expressed and co-exist as heterodimers in uterine and blastocyst tissues. The ISP1-ISP2 enzyme complex shows trypsin-like substrate specificity. In contrast, we found that ISP2, isolated as a 34 kDa monomer from a Pichia pastoris expression system, exhibited a mixed serine proteolytic substrate specificity, as determined by a phage display peptide cleavage approach and verified by the in vitro cleavage of synthetic peptides. Based upon the peptide sequence substrate selectivity, a database search identified many potential ISP2 targets of physiological relevance, including the proteinase activated receptor 2 (PAR2). The in vitro cleavage studies with PAR2-derived peptides confirmed the mixed substrate specificity of ISP2. Treatment of cell lines expressing proteinase-activated receptors (PARs) 1, 2, and 4 with ISP2 prevented receptor activation by either thrombin (PARs 1 and 4) or trypsin (PAR2). The disarming and silencing of PARs by ISP2 may play a role in successful embryo implantation.
a Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Calgary, Calgary, Alberta T2N 4N1, Canada.
Navneet
N
Sharma
Jochen
J
Fahr
Bernard
B
Renaux
Mahmoud
M
Saifeddine
Rajeev
R
Kumar
Sandra
S
Nishikawa
Koichiro
K
Mihara
Rithwik
R
Ramachandran
Morley D
MD
Hollenberg
Derrick E
DE
Rancourt
D000595:Amino Acid Sequence; D000818:Animals; D010064:Embryo Implantation; D005260:Female; D015870:Gene Expression; D005786:Gene Expression Regulation; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D010843:Pichia / Q000235:genetics / Q000378:metabolism; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D059748:Proteolysis; D051381:Rats; D044463:Receptor, PAR-1 / Q000235:genetics / Q000378:metabolism*; D044464:Receptor, PAR-2 / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics*; D013917:Thrombin / Q000378:metabolism; D014357:Trypsin / Q000378:metabolism
D019151:Peptide Library; D010455:Peptides; D020033:Protein Isoforms; D044463:Receptor, PAR-1; D044464:Receptor, PAR-2; D011994:Recombinant Proteins; C436409:Prss29 protein, mouse; D012697:Serine Endopeptidases; D014357:Trypsin; D013917:Thrombin