| Abstract
| - A catalytic DNA-templated reaction of hydrolysis of an ester group in an N-modified peptidenucleic acid, which is activated by a Cu2+ complex−PNA, has been discovered and optimized. Both theester-containing PNA and the metal complex PNA bind neighboring sites on a template DNA. This bringsthe reacting groups (the ester and the Cu2+ complex) in proximity to each other and accelerates thehydrolysis of the ester ∼500 times in comparison with its hydrolysis in the absence of the template. Thehydrolysis reaction provides >102-fold kinetic discrimination between DNAs that are different from eachother at a single nucleotide position. Natural enzyme T4 DNA ligase is slightly less selective. On the basisof this reaction a fully homogeneous and sensitive assay for sequence-specific DNA detection has beendeveloped (10 fmol DNA). Identification of one of four DNAs (variation at one position) can be done in asingle experiment. Since the Cu2+ ion is tightly bound in an associate containing the ester PNA, the metalcomplex PNA, and the template DNA, application of this method in buffers containing other Cu2+-bindingligands, e.g., PCR buffer and physiological buffer, is possible.
|
