title: Computational design of an α-gliadin peptidase.
abstract: The ability to rationally modify enzymes to perform novel chemical transformations is essential for the rapid production of next-generation protein therapeutics. Here we describe the use of chemical principles to identify a naturally occurring acid-active peptidase, and the subsequent use of computational protein design tools to reengineer its specificity toward immunogenic elements found in gluten that are the proposed cause of celiac disease. The engineered enzyme exhibits a k(cat)/K(M) of 568 M(-1) s(-1), representing a 116-fold greater proteolytic activity for a model gluten tetrapeptide than the native template enzyme, as well as an over 800-fold switch in substrate specificity toward immunogenic portions of gluten peptides. The computationally engineered enzyme is resistant to proteolysis by digestive proteases and degrades over 95% of an immunogenic peptide implicated in celiac disease in under an hour. Thus, through identification of a natural enzyme with the pre-existing qualities relevant to an ultimate goal and redefinition of its substrate specificity using computational modeling, we were able to generate an enzyme with potential as a therapeutic for celiac disease.
authors.affiliation: Department of Biochemistry, University of Washington, Seattle, Washington 98195, United States.
authors.forename: Sydney R
authors.initials: SR
authors.lastname: Gordon
authors.forename: Elizabeth J
authors.initials: EJ
authors.lastname: Stanley
authors.forename: Sarah
authors.initials: S
authors.lastname: Wolf
authors.forename: Angus
authors.initials: A
authors.lastname: Toland
authors.forename: Sean J
authors.initials: SJ
authors.lastname: Wu
authors.forename: Daniel
authors.initials: D
authors.lastname: Hadidi
authors.forename: Jeremy H
authors.initials: JH
authors.lastname: Mills
authors.forename: David
authors.initials: D
authors.lastname: Baker
authors.forename: Ingrid Swanson
authors.initials: IS
authors.lastname: Pultz
authors.forename: Justin B
authors.initials: JB
authors.lastname: Siegel
mesh_terms: D000595:Amino Acid Sequence; D005903:Gliadin / Q000737:chemistry*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010447:Peptide Hydrolases / Q000737:chemistry*
chemical_list: D005903:Gliadin; D010447:Peptide Hydrolases