title: Conservation of the extended substrate specificity profiles among homologous granzymes across species.
abstract: Granzymes are structurally related serine proteases involved in cell death and immunity. To date four out of five human granzymes have assigned orthologs in mice; however for granzyme H, no murine ortholog has been suggested and its role in cytotoxicity remains controversial. Here, we demonstrate that, as is the case for granzyme C, human granzyme H is an inefficient cytotoxin that together with their similar pattern of GrB divergence and functional similarity strongly hint to their orthologous relationship. Besides analyzing the substrate specificity profile of granzyme H by substrate phage display, substrate cleavage susceptibility of human granzyme H and mouse granzyme C was assessed on a proteome-wide level. The extended specificity profiles of granzymes C and H (i.e. beyond cleavage positions P4-P4') match those previously observed for granzyme B. We demonstrate conservation of these extended specificity profiles among various granzymes as granzyme B cleavage susceptibility of an otherwise granzyme H/C specific cleavage site can simply be conferred by altering the P1-residue to aspartate, the preferred P1-residue of granzyme B. Our results thus indicate a conserved, but hitherto underappreciated specificity-determining role of extended protease-substrate contacts in steering cleavage susceptibility.
authors.affiliation: Department of Medical Protein Research, VIB, B-9000 Ghent, Belgium;
authors.forename: Kim
authors.initials: K
authors.lastname: Plasman
authors.forename: Sebastian
authors.initials: S
authors.lastname: Maurer-Stroh
authors.forename: Jamshaid
authors.initials: J
authors.lastname: Ahmad
authors.forename: Han
authors.initials: H
authors.lastname: Hao
authors.forename: Dion
authors.initials: D
authors.lastname: Kaiserman
authors.forename: Fernanda L
authors.initials: FL
authors.lastname: Sirota
authors.forename: Veronique
authors.initials: V
authors.lastname: Jonckheere
authors.forename: Phillip I
authors.initials: PI
authors.lastname: Bird
authors.forename: Kris
authors.initials: K
authors.lastname: Gevaert
authors.forename: Petra
authors.initials: P
authors.lastname: Van Damme
mesh_terms: D000818:Animals; D002460:Cell Line; D053804:Granzymes / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D020014:K562 Cells; D051379:Mice; D040901:Proteomics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D011994:Recombinant Proteins; D053804:Granzymes