Dynamic Disorder in Single-Enzyme Experiments: Facts and Artifacts

Abstract

Using a single molecule fluorescence approach, the time series of catalytic events of an enzymatic reaction can be monitored yielding a sequence of fluorescent "on"- and "off"-states. An accurate on/off-assignment is complicated by the intrinsic and extrinsic noise in every single molecule fluorescence experiment. Using simulated data, the performance of the most widely employed binning and thresholding approach was systematically compared to change point analysis. It is shown that the underlying on- and off-histograms as well as the off-autocorrelation are not necessarily extracted from the "signal" buried in noise. The shapes of the on- and off-histograms are affected by artifacts introduced by the analysis procedure and depend on the signal-to-noise ratio and the overall fluorescence intensity. For experimental data where the background Intensity is not constant over time we consider change point analysis to be more accurate. When using change point analysis for data of the enzyme α-chymotrypsin, no characteristics of dynamic disorder were found. In light of these results, dynamic disorder might not be a general characteristic of enzymatic reactions

Non-Markovian properties and multiscale hidden Markovian network buried in single molecule time series

“…37 Recently, it was shown that the existence of dynamic disorder of single enzymatic turnover reactions depends on how one assigns the ON/OFF levels and the most widely used binning and thresholding approach may yield a misleading interpretation. 38 Because some subsequences involving one step transitions in the ON/OFF time series were removed from the analyses, 16 it is desired to confirm how such removals would affect in the analyses based on change point detection method. 36,38 It should also be noted that the SSN scheme groups a set of past subsequences (into a single state) whose transition probability distributions are regarded as identical within an error tolerance determined by the significance level.…”

“…38 Because some subsequences involving one step transitions in the ON/OFF time series were removed from the analyses, 16 it is desired to confirm how such removals would affect in the analyses based on change point detection method. 36,38 It should also be noted that the SSN scheme groups a set of past subsequences (into a single state) whose transition probability distributions are regarded as identical within an error tolerance determined by the significance level. The grouping and resultant states depend on this significance level and even if two past subsequences are grouped into a certain state for a specific significance level, it might not be the case for another significance level.…”

“…The symbolization and the number of symbols depend on the nature of time series, experimental setup, signal-to-noise ratio, and so forth. 23,36,38 The second step is to evaluate the transition probabilities from different subsequences (called past subsequences) to the future symbols, e.g., the transition probability P(s i |s 2 s 1 ) for any symbol s i (i=1,..., the total number of symbols) to appear at a time, say t, following a particular subsequence s 2 s 1 in which one observes s 1 at time t − 1 and s 2 at time t − 2. Similarly, the transition probabilities for all other past subsequences s j s k with past length two will be evaluated.…”

Kinetic Modelling of Enzyme Catalyzed Biotransformation Involving Activations and Inhibitions

“…This phenomenon is called dynamic disorder [120][121][122][123]. Single molecule experiments have shown that the stochastic waiting time of an enzymatic reaction exhibits a distribution of an exponential rise followed by an exponential decay [124,125]. For a single molecule with slowly interconverting conformational states with different k cat and K m , it follows that…”

Order and disorder in irreversible decay processes

“…More recent advances in single-molecule techniques are bringing attention to the disordered decay 7 of structural correlations in biomolecules during electron transfer reactions 8 and enzyme catalyzed reactions. [9][10][11][12] Experimental techniques have also inspired new theoretical approaches to disora) jason.green@umb.edu dered kinetics [13][14][15] in the forced unfolding 16,17 and spontaneous folding 18,19 of biomolecules. Disordered kinetics is also of interest in the charge transfer through molecular assemblies because of their potential use in molecular electronics, solar cells, and artificial photosynthesis.…”