Structure-function investigation of a deoxyribozyme with dual chelatase and peroxidase activities

PS2.M, an 18-nucleotide DNA molecule, has been shown to be a dual enzyme for porphyrin metallation and, when complexed with hemin, for peroxidation. To date, detailed information has not been available on either the actively folded structure of PS2.M or on the contribution of specific nucleotides within it toward the peroxidase activity. Here, we report a variety of experiments that probe the structure and function of PS2.M as well as of a number of point mutants of PS2.M. Based on these experiments, a structural model for the folding of PS2.M and the location of a functionally relevant hemin-binding site are proposed. A key finding is that PS2.M, originally obtained by in vitro selection from a random-sequence DNA library, is uniquely suited for its catalysis of peroxidation; all point mutants examined showed significantly poorer catalytic activity than PS2.M itself.