Dynamics of Nuclear Receptor Helix-12 Switch of Transcription Activation by Modeling Time-Resolved Fluorescence Anisotropy Decays

Batista, Mariana; Martı́nez, Leandro

doi:10.1016/j.bpj.2013.07.032

Cited by 35 publications

(56 citation statements)

References 57 publications

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“…Both experimental and modeling studies are inconsistent with radical repositioning of H12 away from the AF-2 in apo-NRs (53)(54)(55)(56). Rather, subtle local conformational adaptations are observed in H12 as well as other regions within the ligand binding pocket such as the H11-H12 loop, H3, and H5 (56).…”

Section: Discussionmentioning

confidence: 91%

Unexpected Allosteric Network Contributes to LRH-1 Co-regulator Selectivity

Musille

Kossmann

Kohn

et al. 2016

Journal of Biological Chemistry

103

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Phospholipids (PLs) are unusual signaling hormones sensed by the nuclear receptor liver receptor homolog-1 (LRH-1), which has evolved a novel allosteric pathway to support appropriate interaction with co-regulators depending on ligand status. LRH-1 plays an important role in controlling lipid and cholesterol homeostasis and is a potential target for the treatment of metabolic and neoplastic diseases. Although the prospect of modulating LRH-1 via small molecules is exciting, the molecular mechanism linking PL structure to transcriptional co-regulator preference is unknown. Previous studies showed that binding to an activating PL ligand, such as dilauroylphosphatidylcholine, favors LRH-1's interaction with transcriptional coactivators to up-regulate gene expression. Both crystallographic and solution-based structural studies showed that dilauroylphosphatidylcholine binding drives unanticipated structural fluctuations outside of the canonical activation surface in an alternate activation function (AF) region, encompassing the ␤-sheet-H6 region of the protein. However, the mechanism by which dynamics in the alternate AF influences co-regulator selectivity remains elusive. Here, we pair x-ray crystallography with molecular modeling to identify an unexpected allosteric network that traverses the protein ligand binding pocket and links these two elements to dictate selectivity. We show that communication between the alternate AF region and classical AF2 is correlated with the strength of the co-regulator interaction. This work offers the first glimpse into the conformational dynamics that drive this unusual PL-mediated nuclear hormone receptor activation.

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Section: Discussionmentioning

confidence: 91%

Unexpected Allosteric Network Contributes to LRH-1 Co-regulator Selectivity

Musille

Kossmann

Kohn

et al. 2016

Journal of Biological Chemistry

103

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“…1F, Upper). Simultaneous measurements of photon chronic arrival time and delay time along with rotational anisotropy improve the resolution of detection of individual species and provide rich information about the properties of a fluorophore and its surrounding environment (9,(31)(32)(33)(34)(35)(36). In this regard our study is unique and advanced in interrogating the productreleasing mechanism and associated conformational dynamics.…”

Section: Resultsmentioning

confidence: 98%

Probing conformational dynamics of an enzymatic active site by an in situ single fluorogenic probe under piconewton force manipulation

Pal

2016

Proc. Natl. Acad. Sci. U.S.A.

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Unraveling the conformational details of an enzyme during the essential steps of a catalytic reaction (i.e., enzyme-substrate interaction, enzyme-substrate active complex formation, nascent product formation, and product release) is challenging due to the transient nature of intermediate conformational states, conformational fluctuations, and the associated complex dynamics. Here we report our study on the conformational dynamics of horseradish peroxidase using single-molecule multiparameter photon time-stamping spectroscopy with mechanical force manipulation, a newly developed single-molecule fluorescence imaging magnetic tweezers nanoscopic approach. A nascent-formed fluorogenic product molecule serves as a probe, perfectly fitting in the enzymatic reaction active site for probing the enzymatic conformational dynamics. Interestingly, the product releasing dynamics shows the complex conformational behavior with multiple product releasing pathways. However, under magnetic force manipulation, the complex nature of the multiple product releasing pathways disappears and more simplistic conformations of the active site are populated.enzymatic conformational dynamics | magnetic tweezers | mechanical force manipulation | enzymatic product releasing | fluorogenic substrate E nzymes are capable of enhancing biological reaction activity by millions or even billions of times (1). A typical enzymatic turnover cycle involves multiple steps: substrate (S) binding to the active site of the enzyme (E) in forming an enzyme-substrate complex E+S→[E · S]; enzymatic reaction converting substrate to nascent product (P), [E · S]→[E · P]; and product releasing from the enzyme active site, [E · P]→E+P, which completes an enzymatic reaction turnover (2-4). In recent years, it has been widely identified that conformational dynamics plays a crucial role in regulating enzymatic reactions (2, 4-7). The advent of sitespecific mutagenesis, single-molecule (SM) spectroscopic technique, and molecular dynamics simulations have demonstrated complex dynamics involving multiple conformational states during a catalytic cycle (4, 5, 7-13). Fundamentally, not only the enzymatic active site conformational dynamics but also the overall conformational fluctuation dynamics of the protein matrix, providing the required entropy, enthalpy, and corresponding energy landscape, has a profound impact on an enzyme to ultimately possess the power of enhancing reaction rates. Nevertheless, capturing an individual snapshot of each intermediate conformational step remains challenging, mainly due to the transient nature of those intermediate structures, inadequacy of the conventional structure analysis methods to seize those fluctuating structures, and lack of molecular probes that can specifically report the active site environment at each step of an enzymatic reaction. More often than not, the rate-limiting step is the product release from the active site (14). However, a thorough and comprehensive picture of the product release mechanism is still insufficient, mos...

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“…The variability of the molecular environment and its effects on the Trp conformations has been used to study the time‐dependence of fluorescence wavelength and anisotropy decay . Here, different protein and solvent conformations obtained in the simulations are associated with different solvent accessibilities and electrostatic interactions of the indole group.…”

Section: Resultsmentioning

confidence: 99%

Parametric models to compute tryptophan fluorescence wavelengths from classical protein simulations

Lopez

Martı́nez

2018

J Comput Chem

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Fluorescence spectroscopy is an important method to study protein conformational dynamics and solvation structures. Tryptophan (Trp) residues are the most important and practical intrinsic probes for protein fluorescence due to the variability of their fluorescence wavelengths: Trp residues emit in wavelengths ranging from 308 to 360 nm depending on the local molecular environment. Fluorescence involves electronic transitions, thus its computational modeling is a challenging task. We show that it is possible to predict the wavelength of emission of a Trp residue from classical molecular dynamics simulations by computing the solvent-accessible surface area or the electrostatic interaction between the indole group and the rest of the system. Linear parametric models are obtained to predict the maximum emission wavelengths with standard errors of the order 5 nm. In a set of 19 proteins with emission wavelengths ranging from 308 to 352 nm, the best model predicts the maximum wavelength of emission with a standard error of 4.89 nm and a quadratic Pearson correlation coefficient of 0.81. These models can be used for the interpretation of fluorescence spectra of proteins with multiple Trp residues, or for which local Trp environmental variability exists and can be probed by classical molecular dynamics simulations. © 2018 Wiley Periodicals, Inc.

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Dynamics of Nuclear Receptor Helix-12 Switch of Transcription Activation by Modeling Time-Resolved Fluorescence Anisotropy Decays

Cited by 35 publications

References 57 publications

Unexpected Allosteric Network Contributes to LRH-1 Co-regulator Selectivity

Unexpected Allosteric Network Contributes to LRH-1 Co-regulator Selectivity

Probing conformational dynamics of an enzymatic active site by an in situ single fluorogenic probe under piconewton force manipulation

Parametric models to compute tryptophan fluorescence wavelengths from classical protein simulations

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