Rakesh K Mishra
Articles written in Journal of Chemical Sciences
Volume 106 Issue 5 October 1994 pp 1071-1088 Nucleic Acids: Synthesis, Structure and Application
A central strategy for the design of chemical nucleases is presented. This involves the utilization of the α-amino carboxylate unit of
Fluorescence quenching studies with Bz-Trp-Trp-K-Cu-K-Trp-Trp-Bz demonstrated the binding of one ligand at approximately every stretch of 112 bp and approximately a stretch of 80 bp in the presence of salt. The DNA cleaving properties of all the nucleases were demonstrated with pBR 322 and p blue script 11KS using standard protocols. In all cases, covalently closed supercoiled (form I DNA) is converted largely into open circular (form II) suggesting nicking of the single strand at binding sites. Sequence specificity experiments with the nuclease. Bz-Ala-Gly-K-Cu-K-Gly-Ala-Bz in a32P 3’-end labeled 117 bp restriction fragment (Eco RI/Hind III) of pUC-18 showed almost exclusive attacks at thymidylate residues in particular, thymines corresponding to 5’T of the CTAT(3’-5’) box. Whilst the most preferred site of attack is found at T of 3’-ATC-5’ at the trinucleotide level, cleavage studies at low concentration have shown that at pentanucleotide level, the lone sequence 3’-GATCT-5’ (a part of the inverted repeat -GAGATCTC-) is favoured (fragment 92) over the more frequently occurring 3’-TATCT-5’ segment.
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