A ribozyme and a catalytic DNA with peroxidase activity: active sites versus cofactor-binding sites.
BACKGROUND
An 18-nucleotide DNA oligomer, PS2.M, derived using an in vitro selection method was previously reported to bind hemin (Fe(III)-protoporphyrinIX) with submicromolar affinity. The DNA-hemin complex exhibited DNA-enhanced peroxidative activity. PS2. M is guanine-rich and requires potassium ions to fold to its active conformation, consistent with its forming a guanine-quaduplex. In investigating the specific catalytic features of PS2.M we tested the peroxidative properties of its RNA version (rPS2.M) as well as that of an unrelated DNA guanine-quadruplex, OXY4.


RESULTS
The hemin-binding affinity of rPS2.M was found to be 30-fold weaker than that of PS2.M. The UV-visible spectra and kinetics of enzymatic peroxidation of the RNA-hemin complex, however, were nearly identical to those of its DNA counterpart. Both displayed peroxidase activity substantially greater than those of heme proteins such as catalase and Fe(III)-myoglobin. Kinetic analysis suggested that PS2. M and rPS2.M catalyzed the breakdown of the hemin-hydrogen peroxide covalent complex to products. The hemin complex of folded OXY4 (which bound hemin as strongly as did rPS2.M) had a distinct absorption spectrum and only a minor peroxidase activity above the background level.


CONCLUSIONS
The results indicated that it is possible for RNA and DNA of the same sequence to fold to form comparable cofactor-binding sites, and to show comparable catalytic behavior. The results further suggest that only a subset of cofactor-binding sites formed within folded nucleic acids might be able to function as active sites, by providing the appropriate chemical environments for catalysis.
Travascio
P
P
Department of Chemistry, Institute of Molecular Biology & Biochemistry, Simon Fraser University, Burnaby, V5A 1S6, BC Canada.
Bennet
A J
AJ
Wang
D Y
DY
Sen
D
D
D001483:Base Sequence; D001665:Binding Sites; D002384:Catalysis; D021881:DNA, Catalytic; D004277:DNA, Single-Stranded / Q000378:metabolism*; D006427:Hemin / Q000378:metabolism; D007700:Kinetics; D008786:Metmyoglobin / Q000378:metabolism; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000737:chemistry; D009843:Oligoribonucleotides / Q000138:chemical synthesis / Q000737:chemistry; D010544:Peroxidases / Q000378:metabolism*; D016337:RNA, Catalytic / Q000378:metabolism*; D013379:Substrate Specificity
D021881:DNA, Catalytic; D004277:DNA, Single-Stranded; D009838:Oligodeoxyribonucleotides; D009843:Oligoribonucleotides; D016337:RNA, Catalytic; D008786:Metmyoglobin; D006427:Hemin; D010544:Peroxidases