Publication

Forster resonance energy transfer and protein-induced fluorescence enhancement as synergetic multiscale molecular rulers

Ploetz, E., Lerner, E., Husada, F., Roelfs, M., Chung, S., Hohlbein, J., Weiss, S. & Cordes, T., 19-Sep-2016, In : Scientific Reports. 6, 18 p., 33257.

Research output: Contribution to journalArticleAcademicpeer-review

APA

Ploetz, E., Lerner, E., Husada, F., Roelfs, M., Chung, S., Hohlbein, J., ... Cordes, T. (2016). Forster resonance energy transfer and protein-induced fluorescence enhancement as synergetic multiscale molecular rulers. Scientific Reports, 6, [33257]. https://doi.org/10.1038/srep33257

Author

Ploetz, Evelyn ; Lerner, Eitan ; Husada, Florence ; Roelfs, Martin ; Chung, SangYoon ; Hohlbein, Johannes ; Weiss, Shimon ; Cordes, Thorben. / Forster resonance energy transfer and protein-induced fluorescence enhancement as synergetic multiscale molecular rulers. In: Scientific Reports. 2016 ; Vol. 6.

Harvard

Ploetz, E, Lerner, E, Husada, F, Roelfs, M, Chung, S, Hohlbein, J, Weiss, S & Cordes, T 2016, 'Forster resonance energy transfer and protein-induced fluorescence enhancement as synergetic multiscale molecular rulers', Scientific Reports, vol. 6, 33257. https://doi.org/10.1038/srep33257

Standard

Forster resonance energy transfer and protein-induced fluorescence enhancement as synergetic multiscale molecular rulers. / Ploetz, Evelyn; Lerner, Eitan; Husada, Florence; Roelfs, Martin; Chung, SangYoon; Hohlbein, Johannes; Weiss, Shimon; Cordes, Thorben.

In: Scientific Reports, Vol. 6, 33257, 19.09.2016.

Research output: Contribution to journalArticleAcademicpeer-review

Vancouver

Ploetz E, Lerner E, Husada F, Roelfs M, Chung S, Hohlbein J et al. Forster resonance energy transfer and protein-induced fluorescence enhancement as synergetic multiscale molecular rulers. Scientific Reports. 2016 Sep 19;6. 33257. https://doi.org/10.1038/srep33257


BibTeX

@article{daaa224cbeea4a8998926238235efe83,
title = "Forster resonance energy transfer and protein-induced fluorescence enhancement as synergetic multiscale molecular rulers",
abstract = "Advanced microscopy methods allow obtaining information on (dynamic) conformational changes in biomolecules via measuring a single molecular distance in the structure. It is, however, extremely challenging to capture the full depth of a three-dimensional biochemical state, binding-related structural changes or conformational cross-talk in multi-protein complexes using one-dimensional assays. In this paper we address this fundamental problem by extending the standard molecular ruler based on Forster resonance energy transfer (FRET) into a two-dimensional assay via its combination with protein-induced fluorescence enhancement (PIFE). We show that donor brightness (via PIFE) and energy transfer efficiency (via FRET) can simultaneously report on e.g., the conformational state of double stranded DNA (dsDNA) following its interaction with unlabelled proteins (BamHI, EcoRV, and T7 DNA polymerase gp5/trx). The PIFE-FRET assay uses established labelling protocols and single molecule fluorescence detection schemes (alternating-laser excitation, ALEX). Besides quantitative studies of PIFE and FRET ruler characteristics, we outline possible applications of ALEX-based PIFE-FRET for single-molecule studies with diffusing and immobilized molecules. Finally, we study transcription initiation and scrunching of E. coli RNA-polymerase with PIFE-FRET and provide direct evidence for the physical presence and vicinity of the polymerase that causes structural changes and scrunching of the transcriptional DNA bubble.",
keywords = "ALTERNATING-LASER EXCITATION, SINGLE-STRANDED-DNA, PHOTOINDUCED ELECTRON-TRANSFER, RNA-POLYMERASE PROCEEDS, CONFORMATIONAL DYNAMICS, T7 DNA, CORRELATION SPECTROSCOPY, INTRAMOLECULAR PHOTOSTABILIZATION, ABORTIVE INITIATION, STRUCTURAL DYNAMICS",
author = "Evelyn Ploetz and Eitan Lerner and Florence Husada and Martin Roelfs and SangYoon Chung and Johannes Hohlbein and Shimon Weiss and Thorben Cordes",
year = "2016",
month = "9",
day = "19",
doi = "10.1038/srep33257",
language = "English",
volume = "6",
journal = "Scientific Reports",
issn = "2045-2322",
publisher = "Nature Publishing Group",

}

RIS

TY - JOUR

T1 - Forster resonance energy transfer and protein-induced fluorescence enhancement as synergetic multiscale molecular rulers

AU - Ploetz, Evelyn

AU - Lerner, Eitan

AU - Husada, Florence

AU - Roelfs, Martin

AU - Chung, SangYoon

AU - Hohlbein, Johannes

AU - Weiss, Shimon

AU - Cordes, Thorben

PY - 2016/9/19

Y1 - 2016/9/19

N2 - Advanced microscopy methods allow obtaining information on (dynamic) conformational changes in biomolecules via measuring a single molecular distance in the structure. It is, however, extremely challenging to capture the full depth of a three-dimensional biochemical state, binding-related structural changes or conformational cross-talk in multi-protein complexes using one-dimensional assays. In this paper we address this fundamental problem by extending the standard molecular ruler based on Forster resonance energy transfer (FRET) into a two-dimensional assay via its combination with protein-induced fluorescence enhancement (PIFE). We show that donor brightness (via PIFE) and energy transfer efficiency (via FRET) can simultaneously report on e.g., the conformational state of double stranded DNA (dsDNA) following its interaction with unlabelled proteins (BamHI, EcoRV, and T7 DNA polymerase gp5/trx). The PIFE-FRET assay uses established labelling protocols and single molecule fluorescence detection schemes (alternating-laser excitation, ALEX). Besides quantitative studies of PIFE and FRET ruler characteristics, we outline possible applications of ALEX-based PIFE-FRET for single-molecule studies with diffusing and immobilized molecules. Finally, we study transcription initiation and scrunching of E. coli RNA-polymerase with PIFE-FRET and provide direct evidence for the physical presence and vicinity of the polymerase that causes structural changes and scrunching of the transcriptional DNA bubble.

AB - Advanced microscopy methods allow obtaining information on (dynamic) conformational changes in biomolecules via measuring a single molecular distance in the structure. It is, however, extremely challenging to capture the full depth of a three-dimensional biochemical state, binding-related structural changes or conformational cross-talk in multi-protein complexes using one-dimensional assays. In this paper we address this fundamental problem by extending the standard molecular ruler based on Forster resonance energy transfer (FRET) into a two-dimensional assay via its combination with protein-induced fluorescence enhancement (PIFE). We show that donor brightness (via PIFE) and energy transfer efficiency (via FRET) can simultaneously report on e.g., the conformational state of double stranded DNA (dsDNA) following its interaction with unlabelled proteins (BamHI, EcoRV, and T7 DNA polymerase gp5/trx). The PIFE-FRET assay uses established labelling protocols and single molecule fluorescence detection schemes (alternating-laser excitation, ALEX). Besides quantitative studies of PIFE and FRET ruler characteristics, we outline possible applications of ALEX-based PIFE-FRET for single-molecule studies with diffusing and immobilized molecules. Finally, we study transcription initiation and scrunching of E. coli RNA-polymerase with PIFE-FRET and provide direct evidence for the physical presence and vicinity of the polymerase that causes structural changes and scrunching of the transcriptional DNA bubble.

KW - ALTERNATING-LASER EXCITATION

KW - SINGLE-STRANDED-DNA

KW - PHOTOINDUCED ELECTRON-TRANSFER

KW - RNA-POLYMERASE PROCEEDS

KW - CONFORMATIONAL DYNAMICS

KW - T7 DNA

KW - CORRELATION SPECTROSCOPY

KW - INTRAMOLECULAR PHOTOSTABILIZATION

KW - ABORTIVE INITIATION

KW - STRUCTURAL DYNAMICS

U2 - 10.1038/srep33257

DO - 10.1038/srep33257

M3 - Article

VL - 6

JO - Scientific Reports

JF - Scientific Reports

SN - 2045-2322

M1 - 33257

ER -

ID: 35999689