title: Novel protease inhibitors via computational redesign of subtilisin BPN' propeptide.
abstract: The propeptide domain of subtilisin BPN' functions as a molecular chaperone for its cognate protease yet quickly assumes a predominantly unfolded structure following cleavage by the mature protease. In contrast, structural stabilization of the propeptide domain has been proposed to competitively inhibit protease self-cleavage, suggesting the possibility for the generation of novel proteinaceous subtilisin inhibitors. Using a Rosetta fixed backbone design, we have redesigned the subtilisin BPN' propeptide structure to generate synthetic peptide sequences with increased and tunable structural stability. Molecular dynamics simulations provide supporting evidence that the artificial sequences retain structure without its protease cognate unlike the inherently disordered wild-type propeptide. Experimental evaluation of two designer domains by spectroscopic methods verified their structural integrity. Furthermore, the novel propeptide domains were shown to possess significantly enhanced thermostability. Nevertheless, their modest functional performance as protease inhibitors raises doubt that propeptide stability alone is sufficient for effective inhibitor design.
authors.affiliation: Department of Chemistry, Emory University, 1515 Dickey Drive, Atlanta, GA 30322, USA.
authors.forename: Ashley B
authors.initials: AB
authors.lastname: Daugherty
authors.forename: Pravin
authors.initials: P
authors.lastname: Muthu
authors.forename: Stefan
authors.initials: S
authors.lastname: Lutz
mesh_terms: D000595:Amino Acid Sequence; D002942:Circular Dichroism; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011480:Protease Inhibitors / Q000737:chemistry*; D059748:Proteolysis; D017386:Sequence Homology, Amino Acid; D013050:Spectrometry, Fluorescence; D013381:Subtilisins / Q000737:chemistry* / Q000235:genetics
chemical_list: D011480:Protease Inhibitors; D013381:Subtilisins