Nucleotide specificity versus complex heterogeneity in exonuclease activity measurements.
A recent publication reported on measurements of Exonuclease I activity using a real-time fluorescence method that measures the time required by molecules of Exonuclease I to hydrolyze single-stranded DNA that was synthesized to have two fluorescently labeled nucleotides. The observed fluorescence-intensity curves were interpreted as a sign of strong heterogeneity of the activity of Exonuclease I. Here, I propose a different model, which assumes that Exonuclease I activity is nucleotide-dependent, and that a fluorescent label bound to a nucleotide significantly slows its cleavage rate. The presented model fits the observed data equally well, but can be used to make specific predictions upon observable sequence dependence of measured fluorescence-intensity curves.
Enderlein
Jörg
J
Institut für Neurowissenschaft und Biophysik 1, Forschungszentrum Jülich, Jülich, Germany. j.enderlein@fz-juelich.de
D004277:DNA, Single-Stranded / Q000378:metabolism*; D005090:Exodeoxyribonucleases / Q000378:metabolism*; D005456:Fluorescent Dyes / Q000737:chemistry; D008954:Models, Biological; D009711:Nucleotides / Q000378:metabolism*
D004277:DNA, Single-Stranded; D005456:Fluorescent Dyes; D009711:Nucleotides; D005090:Exodeoxyribonucleases; C028450:exodeoxyribonuclease I