Sequence diversity, metal specificity, and catalytic proficiency of metal-dependent phosphorylating DNA enzymes.
Although DNA has not been found responsible for biological catalysis, many artificial DNA enzymes have been created by "in vitro selection." Here we describe a new selection approach to assess the influence of four common divalent metal ions (Ca(2+), Cu(2+), Mg(2+), and Mn(2+)) on sequence diversity, metal specificity, and catalytic proficiency of self-phosphorylating deoxyribozymes. Numerous autocatalytic DNA sequences were isolated, a majority of which were selected using Cu(2+) or Mn(2+) as the divalent metal cofactor. We found that Cu(2+)- and Mn(2+)-derived deoxyribozymes were strictly metal specific, while those selected by Ca(2+) and Mg(2+) were less specific. Further optimization by in vitro evolution resulted in a Mn(2+)-dependent deoxyribozyme with a k(cat) of 2.8 min(-1). Our findings suggest that DNA has sufficient structural diversity to facilitate efficient catalysis using a broad scope of metal cofactor utilizing mechanisms.
Wang
Wei
W
Department of Biochemistry, McMaster University, Hamilton, Ontario, Canada.
Billen
Lieven P
LP
Li
Yingfu
Y
D001483:Base Sequence; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002384:Catalysis / Q000187:drug effects; D002413:Cations, Divalent / Q000378:metabolism / Q000494:pharmacology; D003300:Copper / Q000378:metabolism / Q000494:pharmacology; D021881:DNA, Catalytic / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008274:Magnesium / Q000378:metabolism / Q000494:pharmacology; D008345:Manganese / Q000378:metabolism / Q000494:pharmacology; D019216:Metals, Heavy / Q000378:metabolism* / Q000494:pharmacology; D008969:Molecular Sequence Data; D010766:Phosphorylation / Q000187:drug effects
D002413:Cations, Divalent; D021881:DNA, Catalytic; D019216:Metals, Heavy; D008345:Manganese; D003300:Copper; D008274:Magnesium; D002118:Calcium