Watching proteins fold one molecule at a time

Rhoades, Elizabeth; Gussakovsky, Eugene; Haran, Gilad

doi:10.1073/pnas.2628068100

Cited by 335 publications

(317 citation statements)

References 30 publications

(28 reference statements)

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“…[5][6][7][8][9] In the last two decades, considerable progress has been made in attaining a global understanding of the mechanisms by which proteins fold thanks to breakthroughs in experiments, 10-12 theory, [13][14][15] and computations. [16][17][18][19] Fast folding experiments 4,11,[20][21][22] and single molecule methods [23][24][25] have begun to provide a direct glimpse into the initial stages of protein folding. These experiments show that there is a great diversity in the routes explored during the transitions from unfolded states to the folded state that were unanticipated in ensemble experiments.…”

Section: Introductionmentioning

confidence: 99%

Minimal Models for Proteins and RNA: From Folding to Function

Pincus

Cho

Hyeon

et al. 2008

Progress in Molecular Biology and Translational Science

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We present a panoramic view of the utility of coarse-grained (CG) models to study folding and functions of proteins and RNA. Drawing largely on the methods developed in our group over the last twenty years, we describe a number of key applications ranging from folding of proteins with disulfide bonds to functions of molecular machines. After presenting the theoretical basis that justifies the use of CG models, we explore the biophysical basis for the emergence of a finite number of folds from lattice models. The lattice model simulations of approach to the folded state show that non-native interactions are relevant only early in the folding process -a finding that rationalizes the success of structure-based models that emphasize native interactions. Applications of off-lattice C α and models that explicitly consider side chains (C α -SCM) to folding of β-hairpin and effects of macromolecular crowding are briefly discussed. Successful applications of a new class of off-lattice models, referred to as the Self- We also present two distinct models for RNA, namely, the Three Site Interaction (TIS) model and the SOP model, that probe forced unfolding and force quench refolding of a simple hairpin and Azoarcus ribozyme. The unfolding pathways of Azoarcus ribozyme depend on the loading rate, while constant force and constant loading rate simulations of the hairpin show that both forced-unfolding and force-quench refolding pathways are heterogeneous. The location of the transition state moves as force is varied. The predictions based on the SOP model show that force-induced unfolding pathways of the ribozyme can be dramatically changed by varying the loading rate. We conclude with a discussion of future prospects for the use of coarse-grained models in addressing problems of outstanding interest in biology.

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Section: Introductionmentioning

confidence: 99%

Minimal Models for Proteins and RNA: From Folding to Function

Pincus

Cho

Hyeon

et al. 2008

Progress in Molecular Biology and Translational Science

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“…A promising new approach for anchoring molecules to a surface is to encapsulate them within a lipid vesicle and then tether the vesicle to a solid surface 99,100 (Fig. 2E).…”

Section: Lipid Vesiclesmentioning

confidence: 99%

The importance of surfaces in single-molecule bioscience

2008

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The last ten years have witnessed an explosion of new techniques that can be used to probe the dynamic behavior of individual biological molecules, leading to discoveries that would not have been possible with more traditional biochemical methods. A common feature among these singlemolecule approaches is the need for the biological molecules to be anchored to a solid support surface. This must be done under conditions that minimize nonspecific adsorption without compromising the biological integrity of the sample. In this review we highlight why surface attachments are a critical aspect of many single-molecule studies and we discuss current methods for anchoring biomolecules. Finally, we provide a detailed description of a new method developed by our laboratory for anchoring and organizing hundreds of individual DNA molecules on a surface, allowing "high-throughput" studies of protein-DNA interactions at the single-molecule level.

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“…The vesicle serves as a cage in which the protein is completely solvated, and protein-surface interactions are prevented. Rhoades et al studied folding transitions of single adenylate kinase proteins trapped in glass surface-tethered vesicles (25 ). With FRET, folding transitions appeared in fluorescence time traces as anticorrelated steps of the donor and the acceptor signal of the encapsulated protein prepared at the thermodynamic mid-transition point (Figure 2b).…”

Section: Protein Foldingmentioning

confidence: 99%

Exploring Life by Single-Molecule Fluorescence Spectroscopy

Neuweiler¹,

Sauer²

2005

Anal. Chem.

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onsidering all the effort scientists put into visualizing individual atoms and molecules, it is amazing that it only takes a standard fluorescence microscope with high collection efficiency and suitable filters to detect the fluorescence emission of a single fluorophore. In <20 years, several innovations and technical breakthroughs have brought us to the point where we can study matter on a molecular scale-a skill unimaginable 50 years ago. Parallel to these developments, biochemistry and molecular and cellular biology have grown from infancy into prominent disciplines. New, exciting insights into biomolecular structure, self-assembly, dynamics, and regulation within living systems are results of refined methodological and experimental developments in biochemistry and molecular biology.We can now directly visualize and track individual molecules in their native environment; this allows us to study biological systems on a molecular scale. For example, single-molecule techniques are used to visualize the enzymatic mechanisms of DNA polymerase, which is responsible for the replication of genetic information. DNA polymerase synthesizes a DNA strand by incorporating nucleotides while moving like a locomotive on the complementary DNA template. We no longer calculate incorporation rates by looking at average DNA lengths; rather, we directly track the individual enzyme that is incorporating the nucleotides. In addition, we gain new insights into the working mechanism and performance of the enzyme.To see the difference between single-molecule and ensemble experiments, imagine that you are at a large station observing hundreds of passengers arriving on an unknown number of trains. From such an observation, you cannot answer questions about the individual routes the trains took, how many passengers boarded the train at which stations and at what times, or how many stops each train made. You observe only an average and can conclude only that trains typically transport hundreds of passengers at a time. If single-molecule spectroscopy is used to monitor biological reactions, individual properties can

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Watching proteins fold one molecule at a time

Cited by 335 publications

References 30 publications

Minimal Models for Proteins and RNA: From Folding to Function

Minimal Models for Proteins and RNA: From Folding to Function

The importance of surfaces in single-molecule bioscience

Exploring Life by Single-Molecule Fluorescence Spectroscopy

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