Computational design of a biologically active enzyme.
Rational design of enzymes is a stringent test of our understanding of protein chemistry and has numerous potential applications. Here, we present and experimentally validate the computational design of enzyme activity in proteins of known structure. We have predicted mutations that introduce triose phosphate isomerase activity into ribose-binding protein, a receptor that normally lacks enzyme activity. The resulting designs contain 18 to 22 mutations, exhibit 10(5)- to 10(6)-fold rate enhancements over the uncatalyzed reaction, and are biologically active, in that they support the growth of Escherichia coli under gluconeogenic conditions. The inherent generality of the design method suggests that many enzymes can be designed by this approach.
Dwyer
Mary A
MA
Department of Biochemistry, Duke University Medical Center, Durham, NC 27710, USA.
Looger
Loren L
LL
Hellinga
Homme W
HW
D000465:Algorithms; D001665:Binding Sites; D002384:Catalysis; D020134:Catalytic Domain; D019295:Computational Biology; D003198:Computer Simulation; D004099:Dihydroxyacetone Phosphate / Q000378:metabolism; D019281:Dimerization; D019020:Directed Molecular Evolution; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005986:Glyceraldehyde 3-Phosphate / Q000378:metabolism; D005990:Glycerol / Q000378:metabolism; D006860:Hydrogen Bonding; D007700:Kinetics; D007773:Lactates / Q000378:metabolism; D008024:Ligands; D008958:Models, Molecular; D008968:Molecular Conformation; D009154:Mutation; D033902:Periplasmic Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D015202:Protein Engineering; D011522:Protons; D014305:Triose-Phosphate Isomerase / Q000737:chemistry / Q000378:metabolism
D029968:Escherichia coli Proteins; D007773:Lactates; D008024:Ligands; D033902:Periplasmic Binding Proteins; D011522:Protons; C028003:RbsB protein, E coli; D005986:Glyceraldehyde 3-Phosphate; D004099:Dihydroxyacetone Phosphate; D014305:Triose-Phosphate Isomerase; D005990:Glycerol