Functional compromises among pH tolerance, site specificity, and sequence tolerance for a DNA-hydrolyzing deoxyribozyme.

ABSTRACT: We recently reported the identification by in vitro selection of 10MD5, a deoxyribozyme that requires both Mn2+ and Zn2+ to hydrolyze a single-stranded DNA substrate with formation of 5′-phosphate and 3′-hydroxyl termini. DNA cleavage by 10MD5 proceeds with kobs=2.7 h(−1) and rate enhancement of 10(12) over the uncatalyzed P−O hydrolysis reaction. 10MD5 has a very sharp pH optimum near 7.5, with greatly reduced DNA cleavage rate and yield when the pH is changed by only 0.1 unit in either direction. Here we have optimized 10MD5 by reselection (in vitro evolution), leading to variants with broader pH tolerance, which is important for practical DNA cleavage applications. Because of the extensive Watson−Crick complementarity between deoxyribozyme and substrate, the parent 10MD5 is inherently s

SUBMITTER: Xiao Y 

PROVIDER: S-EPMC3030634 | biostudies-literature | 2010 Nov

REPOSITORIES: biostudies-literature