Single-molecule probing of the conformational homogeneity of the ABC transporter BtuCD

Yang, Min Hsiang; Levanon, Nurit Livnat; Acar, Burcin; Fas, Burcu Aykac; Masrati, Gal; Rose, Jessica; Ben‐Tal, Nir; Haliloğlu, Türkan; Zhao, Yongfang; Lewinson, Oded

doi:10.1038/s41589-018-0088-2

Cited by 54 publications

(87 citation statements)

References 53 publications

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“…Although often hidden from biophysical characterization, they are highly abundant in multidomain proteins such as poly-PDZ proteins 14 , ion channels 52 , ATPases 53 , enzymes 54 , and intrinsically disordered proteins with dynamic supertertiary structures 1 . When identified, these transient and short-lived conformational states could serve as potential targets for drug development and may reveal novel mechanisms of action 55 – 57 .…”

Section: Discussionmentioning

confidence: 99%

Identifying weak interdomain interactions that stabilize the supertertiary structure of the N-terminal tandem PDZ domains of PSD-95

Orozco

Mindlin

et al. 2018

Nat Commun

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Previous studies of the N-terminal PDZ tandem from PSD-95 produced divergent models and failed to identify interdomain contacts stabilizing the structure. We used ensemble and single-molecule FRET along with replica-exchange molecular dynamics to fully characterize the energy landscape. Simulations and experiments identified two conformations: an open-like conformation with a small contact interface stabilized by salt bridges, and a closed-like conformation with a larger contact interface stabilized by surface-exposed hydrophobic residues. Both interfaces were confirmed experimentally. Proximity of interdomain contacts to the binding pockets may explain the observed coupling between conformation and binding. The low-energy barrier between conformations allows submillisecond dynamics, which were time-averaged in previous NMR and FRET studies. Moreover, the small contact interfaces were likely overridden by lattice contacts as crystal structures were rarely sampled in simulations. Our hybrid approach can identify transient interdomain interactions, which are abundant in multidomain proteins yet often obscured by dynamic averaging.

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Section: Discussionmentioning

confidence: 99%

Identifying weak interdomain interactions that stabilize the supertertiary structure of the N-terminal tandem PDZ domains of PSD-95

Orozco

Mindlin

et al. 2018

Nat Commun

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“…Interestingly, many of these studies tended to capture relatively slow conformational dynamics [79,80]. Several experiments focused on ATP-binding cassette (ABC) transporters, which are membrane protein machines that harness ATP energy to move molecules across the cell membrane [81][82][83][84]. ABC transporters are homodimers, each with a substrate-binding domain (SBD) or a separate substrate-binding protein (SBP), a transmembrane domain, and a cytoplasmic nucleotide-binding domain.…”

Section: Implications Of Conformational Dynamics For Proteins Functionmentioning

confidence: 99%

Single-molecule FRET methods to study the dynamics of proteins at work

Mazal

Haran

2019

Current Opinion in Biomedical Engineering

116

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Feynman commented that "Everything that living things do can be understood in terms of the jiggling and wiggling of atoms". Proteins can jiggle and wiggle large structural elements such as domains and subunits as part of their functional cycles. Single-molecule fluorescence resonance energy transfer (smFRET) is an excellent tool to study conformational dynamics and decipher coordinated large-scale motions within proteins. smFRET methods introduced in recent years are geared toward understanding the time scales and amplitudes of function-related motions. This review discusses the methodology for obtaining and analyzing smFRET temporal trajectories that provide direct dynamic information on transitions between conformational states. It also introduces correlation methods that are useful for characterizing intramolecular motions. This arsenal of techniques has been used to study multiple molecular systems, from membrane proteins through molecular chaperones, and we examine some of these studies here. Recent exciting methodological novelties permit revealing very fast, submillisecond dynamics, whose relevance to protein function is yet to be fully grasped.

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“…Tier-0 dynamics were observed and characterized in various settings, e.g., in motor proteins, where they are used in propelling movement along filaments 16 , in the transport of molecules or biopolymers across biological membranes [17][18][19][20][21] , or in the activity of proteins that perform mechanical work. 22 Tier-1 dynamics drive the actions of various signaling proteins for transmission of signals.…”

Section: Introductionmentioning

confidence: 99%

Evolution of structural dynamics in bilobed proteins

Gouridis¹,

Muthahari²,

Boer³

et al. 2020

Preprint

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Novel biophysical tools allow the structural dynamics of proteins, and the regulation of such dynamics by binding partners, to be explored in unprecedented detail. Although this has provided critical insights into protein function, the means by which structural dynamics direct protein evolution remains poorly understood. Here, we investigated how proteins with a bilobed structure, composed of two related domains from the type-II periplasmic binding protein domain family, have undergone divergent evolution leading to modification of their structural dynamics and function. We performed a structural analysis of ~600 bilobed proteins with a common primordial structural core, which we complemented with biophysical studies to explore the structural dynamics of selected examples by single-molecule Foerster resonance energy transfer and Hydrogen-Deuterium exchange mass spectrometry. We show that evolutionary modifications of the structural core, largely at its termini, enables distinct structural dynamics, allowing the diversification of these proteins into transcription factors, enzymes, and extra-cytoplasmic transport-related proteins. Structural embellishments of the core created new interdomain interactions that stabilized structural states, reshaping the active site geometry, and ultimately, altered substrate specificity. Our findings reveal an as yet unrecognized mechanism for the emergence of functional promiscuity during long periods of protein evolution and are applicable to a large number of domain architectures.

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Single-molecule probing of the conformational homogeneity of the ABC transporter BtuCD

Cited by 54 publications

References 53 publications

Identifying weak interdomain interactions that stabilize the supertertiary structure of the N-terminal tandem PDZ domains of PSD-95

Identifying weak interdomain interactions that stabilize the supertertiary structure of the N-terminal tandem PDZ domains of PSD-95

Single-molecule FRET methods to study the dynamics of proteins at work

Evolution of structural dynamics in bilobed proteins

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