title: Protein design algorithms predict viable resistance to an experimental antifolate.
abstract: Methods to accurately predict potential drug target mutations in response to early-stage leads could drive the design of more resilient first generation drug candidates. In this study, a structure-based protein design algorithm (K* in the OSPREY suite) was used to prospectively identify single-nucleotide polymorphisms that confer resistance to an experimental inhibitor effective against dihydrofolate reductase (DHFR) from Staphylococcus aureus. Four of the top-ranked mutations in DHFR were found to be catalytically competent and resistant to the inhibitor. Selection of resistant bacteria in vitro reveals that two of the predicted mutations arise in the background of a compensatory mutation. Using enzyme kinetics, microbiology, and crystal structures of the complexes, we determined the fitness of the mutant enzymes and strains, the structural basis of resistance, and the compensatory relationship of the mutations. To our knowledge, this work illustrates the first application of protein design algorithms to prospectively predict viable resistance mutations that arise in bacteria under antibiotic pressure.
authors.affiliation: Department of Pharmaceutical Sciences, University of Connecticut, Storrs, CT 06269; and.
authors.forename: Stephanie M
authors.initials: SM
authors.lastname: Reeve
authors.affiliation: Departments of Computer Science and.
authors.forename: Pablo
authors.initials: P
authors.lastname: Gainza
authors.affiliation: Department of Pharmaceutical Sciences, University of Connecticut, Storrs, CT 06269; and.
authors.forename: Kathleen M
authors.initials: KM
authors.lastname: Frey
authors.affiliation: Departments of Computer Science and.
authors.forename: Ivelin
authors.initials: I
authors.lastname: Georgiev
authors.affiliation: Departments of Computer Science and Biochemistry, Duke University, Durham, NC 27708 amy.anderson@uconn.edu brd+dhfr14@cs.duke.edu.
authors.forename: Bruce R
authors.initials: BR
authors.lastname: Donald
authors.affiliation: Department of Pharmaceutical Sciences, University of Connecticut, Storrs, CT 06269; and amy.anderson@uconn.edu brd+dhfr14@cs.duke.edu.
authors.forename: Amy C
authors.initials: AC
authors.lastname: Anderson
mesh_terms: D000465:Algorithms; D004351:Drug Resistance / Q000235:genetics; D005493:Folic Acid Antagonists / Q000494:pharmacology*; D020641:Polymorphism, Single Nucleotide; D011506:Proteins / Q000737:chemistry*; D013211:Staphylococcus aureus / Q000201:enzymology; D013762:Tetrahydrofolate Dehydrogenase / Q000187:drug effects
chemical_list: D005493:Folic Acid Antagonists; D011506:Proteins; D013762:Tetrahydrofolate Dehydrogenase
keywords: DHFR; MRSA; antifolate; drug resistance; protein design