Abstract
BackgroundChannel current feature extraction methods, using Hidden Markov Models (HMMs) have been designed for tracking individual-molecule conformational changes. This information is derived from observation of changes in ionic channel current blockade "signal" upon that molecule's interaction with (and occlusion of) a single nanometer-scale channel in a "nanopore detector". In effect, a nanopore detector transduces single molecule events into channel current blockades. HMM analysis tools described are used to help systematically explore DNA dinucleotide flexibility, with particular focus on HIV's highly conserved (and highly flexible/reactive) viral DNA termini. One of the most critical stages in HIV's attack is the binding between viral DNA and the retroviral integrase, which is influenced by the dynamic-coupling induced high flexibility of a CA/TG dinucleotide positioned precisely two base-pairs from the blunt terminus of the duplex viral DNA. This suggests the study of a family of such CA/TG dinucleotide molecules via nanopore measurement and cheminformatics analysis.ResultsHMMs are used for level identification on the current blockades, HMM/EM with boosted variance emissions are used for level projection pre-processing, and time-domain FSAs are used to parse the level-projected waveform for kinetic information. The observed state kinetics of the DNA hairpins containing the CA/TG dinucleotide provides clear evidence for HIV's selection of a peculiarly flexible/interactive DNA terminus.
Highlights
In what follows kinetic feature extraction is done on two types of channel current blockade events: (i) fixed level blockades, and (ii) blockade "spikes"
The blockade level lifetime analysis is primarily Hidden Markov Models (HMMs)-based, where HMM/EM with boosted variance emissions is used for level projection pre-processing, and time-domain Finite State Automaton (FSA) are used to parse the level-projected waveform for kinetic information
Application of the HMM kinetic feature extraction tool permits statistical differences to be discernible between molecules in the study of HIV DNA
Summary
One of the most critical stages in HIV's attack is the binding between viral DNA and the retroviral integrase, which is influenced by the dynamic-coupling induced high flexibility of a CA/TG dinucleotide positioned precisely two base-pairs from the blunt terminus of the duplex viral DNA. This suggests the study of a family of such CA/TG dinucleotide molecules via nanopore measurement and cheminformatics analysis. A test of the hypothesized flexibility/reactivity is sought via analysis of channel current statistics for signs of notably different blockade kinetics between the bluntended HIV DNA conformer and the other blunt-ended hairpins in the CA-step set
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