Dinucleotide junction cleavage versatility of 8-17 deoxyribozyme.
We conducted 16 parallel in vitro selection experiments to isolate catalytic DNAs from a common DNA library for the cleavage of all 16 possible dinucleotide junctions of RNA incorporated into a common DNA/RNA chimeric substrate sequence. We discovered hundreds of sequence variations of the 8-17 deoxyribozyme--an RNA-cleaving catalytic DNA motif previously reported--from nearly all 16 final pools. Sequence analyses identified four absolutely conserved nucleotides in 8-17. Five representative 8-17 variants were tested for substrate cleavage in trans, and together they were able to cleave 14 dinucleotide junctions. New 8-17 variants required Mn2+ to support their broad dinucleotide cleavage capabilities. We hypothesize that 8-17 has a tertiary structure composed of an enzymatic core executing catalysis and a structural facilitator providing structural fine tuning when different dinucleotide junctions are given as cleavage sites.
Cruz
Rani P G
RP
Department of Biochemistry, McMaster University, Hamilton, Canada.
Withers
Johanna B
JB
Li
Yingfu
Y
D002384:Catalysis; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D021881:DNA, Catalytic / Q000737:chemistry / Q000145:classification / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000378:metabolism*; D015226:Dinucleoside Phosphates / Q000737:chemistry / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism; D008345:Manganese / Q000378:metabolism; D008954:Models, Biological; D015394:Molecular Structure; D012313:RNA / Q000737:chemistry / Q000378:metabolism; D016337:RNA, Catalytic / Q000378:metabolism; D017422:Sequence Analysis, DNA; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D021881:DNA, Catalytic; D004277:DNA, Single-Stranded; D015226:Dinucleoside Phosphates; D016337:RNA, Catalytic; C407482:RNA-cleaving DNA 8-17; D008345:Manganese; D012313:RNA; D004247:DNA; D008274:Magnesium