scholarly journals A new view of protein synthesis: Mapping the free energy landscape of the ribosome using single-molecule FRET

Biopolymers ◽  
2008 ◽  
Vol 89 (7) ◽  
pp. 565-577 ◽  
Author(s):  
James B. Munro ◽  
Andrea Vaiana ◽  
Kevin Y. Sanbonmatsu ◽  
Scott C. Blanchard
Keyword(s):  
Free Energy ◽  
Protein Synthesis ◽  
Single Molecule ◽  
Energy Landscape ◽  
Free Energy Landscape ◽  
Single Molecule Fret ◽  
New View

scholarly journals Slow domain reconfiguration causes power-law kinetics in a two-state enzyme

2018 ◽  
Vol 115 (3) ◽  
pp. 513-518 ◽  
Author(s):  
Iris Grossman-Haham ◽  
Gabriel Rosenblum ◽  
Trishool Namani ◽  
Hagen Hofmann
Keyword(s):  
Free Energy ◽  
Single Molecule ◽  
Power Law ◽  
Energy Landscape ◽  
Rate Equations ◽  
Free Energy Landscape ◽  
Closed Domain ◽  
Single Molecule Fret ◽  
Power Law Kinetics ◽  
Interdomain Interactions

Protein dynamics are typically captured well by rate equations that predict exponential decays for two-state reactions. Here, we describe a remarkable exception. The electron-transfer enzyme quiescin sulfhydryl oxidase (QSOX), a natural fusion of two functionally distinct domains, switches between open- and closed-domain arrangements with apparent power-law kinetics. Using single-molecule FRET experiments on time scales from nanoseconds to milliseconds, we show that the unusual open-close kinetics results from slow sampling of an ensemble of disordered domain orientations. While substrate accelerates the kinetics, thus suggesting a substrate-induced switch to an alternative free energy landscape of the enzyme, the power-law behavior is also preserved upon electron load. Our results show that the slow sampling of open conformers is caused by a variety of interdomain interactions that imply a rugged free energy landscape, thus providing a generic mechanism for dynamic disorder in multidomain enzymes.

Single-Molecule FRET Studies of Counterion Effects on the Free Energy Landscape of Human Mitochondrial Lysine tRNA

Biochemistry ◽  
2011 ◽  
Vol 50 (15) ◽  
pp. 3107-3115 ◽  
Author(s):  
Kirsten Dammertz ◽  
Martin Hengesbach ◽  
Mark Helm ◽  
G. Ulrich Nienhaus ◽  
Andrei Yu. Kobitski
Keyword(s):  
Free Energy ◽  
Single Molecule ◽  
Energy Landscape ◽  
Free Energy Landscape ◽  
Single Molecule Fret ◽  
Lysine Trna

scholarly journals Cation-induced kinetic heterogeneity of the intron–exon recognition in single group II introns

2015 ◽  
Vol 112 (11) ◽  
pp. 3403-3408 ◽  
Author(s):  
Danny Kowerko ◽  
Sebastian L. B. König ◽  
Miriam Skilandat ◽  
Daniela Kruschel ◽  
Mélodie C. A. S. Hadzic ◽  
...  
Keyword(s):  
Free Energy ◽  
Single Molecule ◽  
Binding Sites ◽  
Tertiary Structure ◽  
Energy Landscape ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Free Energy Landscape ◽  
Rna Tertiary Structure ◽  
Kinetic Heterogeneity

RNA is commonly believed to undergo a number of sequential folding steps before reaching its functional fold, i.e., the global minimum in the free energy landscape. However, there is accumulating evidence that several functional conformations are often in coexistence, corresponding to multiple (local) minima in the folding landscape. Here we use the 5′-exon–intron recognition duplex of a self-splicing ribozyme as a model system to study the influence of Mg2+ and Ca2+ on RNA tertiary structure formation. Bulk and single-molecule spectroscopy reveal that near-physiological M2+ concentrations strongly promote interstrand association. Moreover, the presence of M2+ leads to pronounced kinetic heterogeneity, suggesting the coexistence of multiple docked and undocked RNA conformations. Heterogeneity is found to decrease at saturating M2+ concentrations. Using NMR, we locate specific Mg2+ binding pockets and quantify their affinity toward Mg2+. Mg2+ pulse experiments show that M2+ exchange occurs on the timescale of seconds. This unprecedented combination of NMR and single-molecule Förster resonance energy transfer demonstrates for the first time to our knowledge that a rugged free energy landscape coincides with incomplete occupation of specific M2+ binding sites at near-physiological M2+ concentrations. Unconventional kinetics in nucleic acid folding frequently encountered in single-molecule experiments are therefore likely to originate from a spectrum of conformations that differ in the occupation of M2+ binding sites.

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