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Explore the nuances of enzyme reaction kinetics on different time scales, from atto-seconds to seconds. Delve into the challenges and intricacies of studying single proteins at work, uncovering insights beyond ensemble measurements. Learn about early breakthroughs in single protein research and the correlation of waiting times in Michaelis-Menten kinetics. Discover new tools and perspectives in studying molecular turnover and reaction trajectories, shedding light on the dynamic world of enzymes at play.
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k1 k2 k3 E•S E0 + P k-1 E0 E Enzyme reaction kinetics E + S
k2[E0][S] = [S] + KM d[P] k-1 + k2 dt KM= k1 Enzyme reaction kinetics = v = k2[E•S]
atto-second femto-second pico-second nano-second micro-second milli-second second Molecular Time scales 10-18 s 10-15 s 10-12 s 10-9 s 10-6 s 10-3 s 1 s
The challenge of one Enzymes are over a million times smaller than a honey bee!
Ion channels Early single protein measurements (1970’s) Neher & Sakmann, Nobel prize medicine 1991
Early optical attempts absorption
Focus on one fluorescence
Ribozyme X. Zhang et al., Science 296, 1473 (2002)
Fluorescence decay rate: Fluctuations: ET model Flavin:NADH oxidoreductase (Fre)
Lifetime H. Yang et al., Science 302, 262 (2003)
waiting time probability density of mean waiting time for Michaelis-Menten kinetics waiting time correlation function intensity correlation function New tools
Single molecule turnovers H.P. Lu et al., Science 282, 1877 (2002)
-galactosidase -galactosidase
Single molecule assay English et al., Nat. Chem. Biol. 2, 87 (2006)
Intensity correlation fluctuation of k2
Enzymes fluctuate on a broad range of time scales Reaction kinetics are dispersed, only the average is measured in ensembles New lessons learned