TY - JOUR
T1 - Systematic evaluation of the dependence of deoxyribozyme catalysis on random region length
AU - Velez, Tania E.
AU - Singh, Jaydeep
AU - Xiao, Ying
AU - Allen, Emily C.
AU - Wong, On Yi
AU - Chandra, Madhavaiah
AU - Kwon, Sarah C.
AU - Silverman, Scott K.
PY - 2012/12/10
Y1 - 2012/12/10
N2 - Functional nucleic acids are DNA and RNA aptamers that bind targets, or they are deoxyribozymes and ribozymes that have catalytic activity. These functional DNA and RNA sequences can be identified from random-sequence pools by in vitro selection, which requires choosing the length of the random region. Shorter random regions allow more complete coverage of sequence space but may not permit the structural complexity necessary for binding or catalysis. In contrast, longer random regions are sampled incompletely but may allow adoption of more complicated structures that enable function. In this study, we systematically examined random region length (N20 through N 60) for two particular deoxyribozyme catalytic activities, DNA cleavage and tyrosine-RNA nucleopeptide linkage formation. For both activities, we previously identified deoxyribozymes using only N40 regions. In the case of DNA cleavage, here we found that shorter N20 and N 30 regions allowed robust catalytic function, either by DNA hydrolysis or by DNA deglycosylation and strand scission via Β-elimination, whereas longer N50 and N60 regions did not lead to catalytically active DNA sequences. Follow-up selections with N20, N30, and N40 regions revealed an interesting interplay of metal ion cofactors and random region length. Separately, for Tyr-RNA linkage formation, N30 and N60 regions provided catalytically active sequences, whereas N20 was unsuccessful, and the N 40 deoxyribozymes were functionally superior (in terms of rate and yield) to N30 and N60. Collectively, the results indicate that with future in vitro selection experiments for DNA and RNA catalysts, and by extension for aptamers, random region length should be an important experimental variable.
AB - Functional nucleic acids are DNA and RNA aptamers that bind targets, or they are deoxyribozymes and ribozymes that have catalytic activity. These functional DNA and RNA sequences can be identified from random-sequence pools by in vitro selection, which requires choosing the length of the random region. Shorter random regions allow more complete coverage of sequence space but may not permit the structural complexity necessary for binding or catalysis. In contrast, longer random regions are sampled incompletely but may allow adoption of more complicated structures that enable function. In this study, we systematically examined random region length (N20 through N 60) for two particular deoxyribozyme catalytic activities, DNA cleavage and tyrosine-RNA nucleopeptide linkage formation. For both activities, we previously identified deoxyribozymes using only N40 regions. In the case of DNA cleavage, here we found that shorter N20 and N 30 regions allowed robust catalytic function, either by DNA hydrolysis or by DNA deglycosylation and strand scission via Β-elimination, whereas longer N50 and N60 regions did not lead to catalytically active DNA sequences. Follow-up selections with N20, N30, and N40 regions revealed an interesting interplay of metal ion cofactors and random region length. Separately, for Tyr-RNA linkage formation, N30 and N60 regions provided catalytically active sequences, whereas N20 was unsuccessful, and the N 40 deoxyribozymes were functionally superior (in terms of rate and yield) to N30 and N60. Collectively, the results indicate that with future in vitro selection experiments for DNA and RNA catalysts, and by extension for aptamers, random region length should be an important experimental variable.
KW - DNA and RNA aptamers
KW - DNA catalysts
KW - RNA catalysts
KW - deoxyribozyme catalysis
KW - nucleic acids
KW - random region length
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U2 - 10.1021/co300111f
DO - 10.1021/co300111f
M3 - Article
C2 - 23088677
AN - SCOPUS:84870937297
SN - 2156-8952
VL - 14
SP - 680
EP - 687
JO - ACS Combinatorial Science
JF - ACS Combinatorial Science
IS - 12
ER -