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Søren Vrønning Hoffmann

Coupled Binding and Helix Formation Monitored by Synchrotron-Radiation Circular Dichroism

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Coupled Binding and Helix Formation Monitored by Synchrotron-Radiation Circular Dichroism. / Karlsson, Elin; Andersson, Eva; Jones, Nykola C.; Hoffmann, Søren Vrønning; Jemth, Per; Kjærgaard, Magnus.

In: Biophysical Journal, Vol. 117, No. 4, 20.08.2019, p. 729-742.

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review

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Author

Karlsson, Elin ; Andersson, Eva ; Jones, Nykola C. ; Hoffmann, Søren Vrønning ; Jemth, Per ; Kjærgaard, Magnus. / Coupled Binding and Helix Formation Monitored by Synchrotron-Radiation Circular Dichroism. In: Biophysical Journal. 2019 ; Vol. 117, No. 4. pp. 729-742.

Bibtex

@article{cba2d85b05494085a96c2c8840b08e56,
title = "Coupled Binding and Helix Formation Monitored by Synchrotron-Radiation Circular Dichroism",
abstract = "Intrinsically disordered proteins organize interaction networks in the cell in many regulation and signaling processes. These proteins often gain structure upon binding to their target proteins in multistep reactions involving the formation of both secondary and tertiary structure. To understand the interactions of disordered proteins, we need to understand the mechanisms of these coupled folding and binding reactions. We studied helix formation in the binding of the molten globule-like nuclear coactivator binding domain and the disordered interaction domain from activator of thyroid hormone and retinoid receptors. We demonstrate that helix formation in a rapid binding reaction can be followed by stopped-flow synchrotron-radiation circular dichroism (CD) spectroscopy and describe the design of such a beamline. Fluorescence-monitored binding experiments of activator of thyroid hormone and retinoid receptors and nuclear coactivator binding domain display several kinetic phases, including one concentration-independent phase, which is consistent with an intermediate stabilized at high ionic strength. Time-resolved CD experiments show that almost all helicity is formed upon initial association of the proteins or separated from the encounter complex by only a small energy barrier. Through simulation of mechanistic models, we show that the intermediate observed at high ionic strength likely involves a structural rearrangement with minor overall changes in helicity. Our experiments provide a benchmark for simulations of coupled binding reactions and demonstrate the feasibility of using synchrotron-radiation CD for mechanistic studies of protein-protein interactions.",
author = "Elin Karlsson and Eva Andersson and Jones, {Nykola C.} and Hoffmann, {S{\o}ren Vr{\o}nning} and Per Jemth and Magnus Kj{\ae}rgaard",
year = "2019",
month = aug,
day = "20",
doi = "10.1016/j.bpj.2019.07.014",
language = "English",
volume = "117",
pages = "729--742",
journal = "Biophysical Journal",
issn = "0006-3495",
publisher = "Cell Press",
number = "4",

}

RIS

TY - JOUR

T1 - Coupled Binding and Helix Formation Monitored by Synchrotron-Radiation Circular Dichroism

AU - Karlsson, Elin

AU - Andersson, Eva

AU - Jones, Nykola C.

AU - Hoffmann, Søren Vrønning

AU - Jemth, Per

AU - Kjærgaard, Magnus

PY - 2019/8/20

Y1 - 2019/8/20

N2 - Intrinsically disordered proteins organize interaction networks in the cell in many regulation and signaling processes. These proteins often gain structure upon binding to their target proteins in multistep reactions involving the formation of both secondary and tertiary structure. To understand the interactions of disordered proteins, we need to understand the mechanisms of these coupled folding and binding reactions. We studied helix formation in the binding of the molten globule-like nuclear coactivator binding domain and the disordered interaction domain from activator of thyroid hormone and retinoid receptors. We demonstrate that helix formation in a rapid binding reaction can be followed by stopped-flow synchrotron-radiation circular dichroism (CD) spectroscopy and describe the design of such a beamline. Fluorescence-monitored binding experiments of activator of thyroid hormone and retinoid receptors and nuclear coactivator binding domain display several kinetic phases, including one concentration-independent phase, which is consistent with an intermediate stabilized at high ionic strength. Time-resolved CD experiments show that almost all helicity is formed upon initial association of the proteins or separated from the encounter complex by only a small energy barrier. Through simulation of mechanistic models, we show that the intermediate observed at high ionic strength likely involves a structural rearrangement with minor overall changes in helicity. Our experiments provide a benchmark for simulations of coupled binding reactions and demonstrate the feasibility of using synchrotron-radiation CD for mechanistic studies of protein-protein interactions.

AB - Intrinsically disordered proteins organize interaction networks in the cell in many regulation and signaling processes. These proteins often gain structure upon binding to their target proteins in multistep reactions involving the formation of both secondary and tertiary structure. To understand the interactions of disordered proteins, we need to understand the mechanisms of these coupled folding and binding reactions. We studied helix formation in the binding of the molten globule-like nuclear coactivator binding domain and the disordered interaction domain from activator of thyroid hormone and retinoid receptors. We demonstrate that helix formation in a rapid binding reaction can be followed by stopped-flow synchrotron-radiation circular dichroism (CD) spectroscopy and describe the design of such a beamline. Fluorescence-monitored binding experiments of activator of thyroid hormone and retinoid receptors and nuclear coactivator binding domain display several kinetic phases, including one concentration-independent phase, which is consistent with an intermediate stabilized at high ionic strength. Time-resolved CD experiments show that almost all helicity is formed upon initial association of the proteins or separated from the encounter complex by only a small energy barrier. Through simulation of mechanistic models, we show that the intermediate observed at high ionic strength likely involves a structural rearrangement with minor overall changes in helicity. Our experiments provide a benchmark for simulations of coupled binding reactions and demonstrate the feasibility of using synchrotron-radiation CD for mechanistic studies of protein-protein interactions.

UR - http://www.scopus.com/inward/record.url?scp=85069975666&partnerID=8YFLogxK

U2 - 10.1016/j.bpj.2019.07.014

DO - 10.1016/j.bpj.2019.07.014

M3 - Journal article

C2 - 31378314

VL - 117

SP - 729

EP - 742

JO - Biophysical Journal

JF - Biophysical Journal

SN - 0006-3495

IS - 4

ER -