DNAzyme footprinting: detecting protein-aptamer complexation on surfaces by blocking DNAzyme cleavage activity.

ABSTRACT: A novel method to quantitatively measure the binding of proteins to single-stranded DNA (ssDNA) aptamers that employs the inhibition of the DNAzyme hydrolysis of aptamer monolayers is described. A 28-base DNAzyme was designed to specifically bind to and cleave a 29-base ssDNA sequence that can fold into a G-quartet aptamer and bind the protein thrombin. The binding strength of the DNAzyme to the aptamer sequence was designed to be less than the binding strength of the thrombin to the aptamer (ΔG° = -43.1 and -51.8 kJ/mol, respectively). Formation of the thrombin-aptamer complex was found to block DNAzyme cleavage activity both in solution and in an ssDNA aptamer monolayer. We denote this method for detecting protein-aptamer complexation as "DNAzyme footprinting" in analogy to the process o

SUBMITTER: Chen Y 

PROVIDER: S-EPMC3576845 | biostudies-literature | 2013 Feb

REPOSITORIES: biostudies-literature