Key result
M1GS ribozymes successfully cleave HCMV UL54 RNA in vitro at an optimal 2:1 ratio.
M1GS ribozymes can be engineered to effectively cleave HCMV UL54 RNA segments in vitro, supporting their potential use as antisense molecules to silence HCMV mRNA.
Does not support clinical use of M1GS ribozymes for HCMV; leaves open therapeutic potential pending in vivo studies.
Seven sequence-specific ribozymes (M1GS RNAs) derived in vitro from the catalytic RNA subunit of Escherichia coli RNase P and targeting the mRNAs transcribed by the UL54 gene encoding the DNA polymerase of human cytomegalovirus were screened from 11 ribozymes that were designed based on four rules: (1) the NCCA-3′terminal must be unpaired with the substrate; (2) the guide sequence (GS) must be at least 12 nt in length; (3) the eighth nucleotide must be U, counting from the site –1; and (4) around the cleavage site, the sites –1/+1/+2 must be U/G/C or C/G/C. Further investigation of the factors affecting the cleavage effect and the optimal ratio for M1GS/substrate was carried out. It was determined that the optimal ratio for M1GS/substrate was 2:1 and too much M1GS led to substrate degrading. As indicated above, several M1GS that cleaved HCMV UL54 RNA segments in vitro were successfully designed and constructed. Our studies support the use of ribozyme M1GS as antisense molecules to silence HCMV mRNA in vitro, and using the selection procedure as a general approach for the engineering of RNase P ribozymes. FullText for HTML: https://doi.org/10.1093/abbs/37.3.210
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SU et al. (2005) studied Human cytomegalovirus (HCMV). M1GS ribozymes targeting HCMV UL54 RNA segments was evaluated on Cleavage of HCMV UL54 RNA segments in vitro. M1GS ribozymes targeting HCMV UL54 RNA segments successfully cleaved the RNA in vitro, with an optimal M1GS/substrate ratio of 2:1.
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