skip to main content
DOE PAGES title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Repurposing a photosynthetic antenna protein as a super-resolution microscopy label

Abstract

Techniques such as Stochastic Optical Reconstruction Microscopy (STORM) and Structured Illumination Microscopy (SIM) have increased the achievable resolution of optical imaging, but few fluorescent proteins are suitable for super-resolution microscopy, particularly in the far-red and near-infrared emission range. Here we demonstrate the applicability of CpcA, a subunit of the photosynthetic antenna complex in cyanobacteria, for STORM and SIM imaging. The periodicity and width of fabricated nanoarrays of CpcA, with a covalently attached phycoerythrobilin (PEB) or phycocyanobilin (PCB) chromophore, matched the lines in reconstructed STORM images. SIM and STORM reconstructions of Escherichia coli cells harbouring CpcA-labelled cytochrome bd 1 ubiquinol oxidase in the cytoplasmic membrane show that CpcA-PEB and CpcA-PCB are suitable for super-resolution imaging in vivo. The stability, ease of production, small size and brightness of CpcA-PEB and CpcA-PCB demonstrate the potential of this largely unexplored protein family as novel probes for super-resolution microscopy.

Authors:
ORCiD logo [1]; ORCiD logo [1];  [2];  [1];  [2];  [1];  [1]; ORCiD logo [2];  [3];  [1];  [2]; ORCiD logo [1]
  1. Univ. of Sheffield (United Kingdom)
  2. Washington Univ., St. Louis, MO (United States)
  3. Pennsylvania State Univ., University Park, PA (United States)
Publication Date:
Research Org.:
Washington Univ., St. Louis, MO (United States); Energy Frontier Research Centers (EFRC) (United States). Photosynthetic Antenna Research Center (PARC)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22)
OSTI Identifier:
1500070
Grant/Contract Number:  
SC0001035
Resource Type:
Accepted Manuscript
Journal Name:
Scientific Reports
Additional Journal Information:
Journal Volume: 7; Journal Issue: 1; Journal ID: ISSN 2045-2322
Publisher:
Nature Publishing Group
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Barnett, Samuel F. H., Hitchcock, Andrew, Mandal, Amit K., Vasilev, Cvetelin, Yuen, Jonathan M., Morby, James, Brindley, Amanda A., Niedzwiedzki, Dariusz M., Bryant, Donald A., Cadby, Ashley J., Holten, Dewey, and Hunter, C. Neil. Repurposing a photosynthetic antenna protein as a super-resolution microscopy label. United States: N. p., 2017. Web. doi:10.1038/s41598-017-16834-z.
Barnett, Samuel F. H., Hitchcock, Andrew, Mandal, Amit K., Vasilev, Cvetelin, Yuen, Jonathan M., Morby, James, Brindley, Amanda A., Niedzwiedzki, Dariusz M., Bryant, Donald A., Cadby, Ashley J., Holten, Dewey, & Hunter, C. Neil. Repurposing a photosynthetic antenna protein as a super-resolution microscopy label. United States. doi:10.1038/s41598-017-16834-z.
Barnett, Samuel F. H., Hitchcock, Andrew, Mandal, Amit K., Vasilev, Cvetelin, Yuen, Jonathan M., Morby, James, Brindley, Amanda A., Niedzwiedzki, Dariusz M., Bryant, Donald A., Cadby, Ashley J., Holten, Dewey, and Hunter, C. Neil. Fri . "Repurposing a photosynthetic antenna protein as a super-resolution microscopy label". United States. doi:10.1038/s41598-017-16834-z. https://www.osti.gov/servlets/purl/1500070.
@article{osti_1500070,
title = {Repurposing a photosynthetic antenna protein as a super-resolution microscopy label},
author = {Barnett, Samuel F. H. and Hitchcock, Andrew and Mandal, Amit K. and Vasilev, Cvetelin and Yuen, Jonathan M. and Morby, James and Brindley, Amanda A. and Niedzwiedzki, Dariusz M. and Bryant, Donald A. and Cadby, Ashley J. and Holten, Dewey and Hunter, C. Neil},
abstractNote = {Techniques such as Stochastic Optical Reconstruction Microscopy (STORM) and Structured Illumination Microscopy (SIM) have increased the achievable resolution of optical imaging, but few fluorescent proteins are suitable for super-resolution microscopy, particularly in the far-red and near-infrared emission range. Here we demonstrate the applicability of CpcA, a subunit of the photosynthetic antenna complex in cyanobacteria, for STORM and SIM imaging. The periodicity and width of fabricated nanoarrays of CpcA, with a covalently attached phycoerythrobilin (PEB) or phycocyanobilin (PCB) chromophore, matched the lines in reconstructed STORM images. SIM and STORM reconstructions of Escherichia coli cells harbouring CpcA-labelled cytochrome bd 1 ubiquinol oxidase in the cytoplasmic membrane show that CpcA-PEB and CpcA-PCB are suitable for super-resolution imaging in vivo. The stability, ease of production, small size and brightness of CpcA-PEB and CpcA-PCB demonstrate the potential of this largely unexplored protein family as novel probes for super-resolution microscopy.},
doi = {10.1038/s41598-017-16834-z},
journal = {Scientific Reports},
number = 1,
volume = 7,
place = {United States},
year = {2017},
month = {12}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record

Figures / Tables:

Figure 1 Figure 1: Photophysical analysis of purified CpcA-PCB and CpcA-PEB. (a) Room temperature absorption and fluorescence emission spectra, normalised for comparison. The same fluorescence spectra were observed for a number of different excitation wavelengths for CpcA-PCB (500, 525, 550, 572 and 625 nm) and CpcA-PEB (490, 510, 525 and 557 nm).more » (b) Fluorescence decay profiles (solid blue and red circles) and dual-exponential fits (solid blue or red lines) of CpcA-PCB using excitation at 582 nm and detection at 644 nm and CpcA-PEB using excitation at 530 nm and detection at 568 nm; the time constants indicated are of the dominant, longer component. The open black circles give the instrument response function, which is approximatly a Gaussian with a full width at half maximum of 200 ps. (c) Representative time profiles (circles) and fits for decay of ground state bleaching (solid lines) or excited state absorption (dashed lines) from the transient absorption data depicted in (d). (d) Time-resolved absorption difference spectra using 100-fs excitation flashes at 590 nm for CpcA-PCB or 510 nm for CpcA-PEB. The data in the region of each spectrum that contains scattered excitation light has been removed.« less

Save / Share:

Works referenced in this record:

Labeling of cytosine residues with biliproteins for use as fluorescent DNA probes
journal, September 2002


Biliproteins and their Applications in Bioimaging
journal, January 2015


Single-molecule in vivo imaging of bacterial respiratory complexes indicates delocalized oxidative phosphorylation
journal, June 2014

  • Llorente-Garcia, Isabel; Lenn, Tchern; Erhardt, Heiko
  • Biochimica et Biophysica Acta (BBA) - Bioenergetics, Vol. 1837, Issue 6
  • DOI: 10.1016/j.bbabio.2014.01.020

Characterization of red-shifted phycobilisomes isolated from the chlorophyll f -containing cyanobacterium Halomicronema hongdechloris
journal, January 2016

  • Li, Yaqiong; Lin, Yuankui; Garvey, Christopher J.
  • Biochimica et Biophysica Acta (BBA) - Bioenergetics, Vol. 1857, Issue 1
  • DOI: 10.1016/j.bbabio.2015.10.009

Synthesis of phycocyanobilin in mammalian cells
journal, January 2013

  • Müller, Konrad; Engesser, Raphael; Timmer, Jens
  • Chemical Communications, Vol. 49, Issue 79
  • DOI: 10.1039/c3cc45065a

Sample drift correction in 3D fluorescence photoactivation localization microscopy
journal, January 2011

  • Mlodzianoski, Michael J.; Schreiner, John M.; Callahan, Steven P.
  • Optics Express, Vol. 19, Issue 16
  • DOI: 10.1364/OE.19.015009

Imaging Intracellular Fluorescent Proteins at Nanometer Resolution
journal, September 2006


Three-Dimensional Resolution Doubling in Wide-Field Fluorescence Microscopy by Structured Illumination
journal, June 2008


Heme oxygenase 1 is required for mammalian iron reutilization
journal, September 1997

  • Poss, K. D.; Tonegawa, S.
  • Proceedings of the National Academy of Sciences, Vol. 94, Issue 20
  • DOI: 10.1073/pnas.94.20.10919

Extensive remodeling of a cyanobacterial photosynthetic apparatus in far-red light
journal, August 2014


Cyan and Yellow Super Fluorescent Proteins with Improved Brightness, Protein Folding, and FRET Förster Radius ,
journal, May 2006

  • Kremers, Gert-Jan; Goedhart, Joachim; van Munster, Erik B.
  • Biochemistry, Vol. 45, Issue 21
  • DOI: 10.1021/bi0516273

Far-red fluorescent tags for protein imaging in living tissues
journal, February 2009

  • Shcherbo, Dmitry; Murphy, Christopher S.; Ermakova, Galina V.
  • Biochemical Journal, Vol. 418, Issue 3
  • DOI: 10.1042/BJ20081949

Photophysical Properties and Electronic Structure of Porphyrins Bearing Zero to Four meso -Phenyl Substituents: New Insights into Seemingly Well Understood Tetrapyrroles
journal, December 2016

  • Mandal, Amit Kumar; Taniguchi, Masahiko; Diers, James R.
  • The Journal of Physical Chemistry A, Vol. 120, Issue 49
  • DOI: 10.1021/acs.jpca.6b09483

Attachment of Noncognate Chromophores to CpcA of Synechocystis sp. PCC 6803 and Synechococcus sp. PCC 7002 by Heterologous Expression in Escherichia coli
journal, June 2011

  • Alvey, Richard M.; Biswas, Avijit; Schluchter, Wendy M.
  • Biochemistry, Vol. 50, Issue 22
  • DOI: 10.1021/bi200307s

Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM)
journal, August 2006

  • Rust, Michael J.; Bates, Mark; Zhuang, Xiaowei
  • Nature Methods, Vol. 3, Issue 10
  • DOI: 10.1038/nmeth929

ThunderSTORM: a comprehensive ImageJ plug-in for PALM and STORM data analysis and super-resolution imaging
journal, May 2014


Occurrence of Far-Red Light Photoacclimation (FaRLiP) in Diverse Cyanobacteria
journal, December 2014


Clustering and dynamics of cytochrome bd -I complexes in the Escherichia coli plasma membrane in vivo
journal, December 2008


SIMcheck: a Toolbox for Successful Super-resolution Structured Illumination Microscopy
journal, November 2015

  • Ball, Graeme; Demmerle, Justin; Kaufmann, Rainer
  • Scientific Reports, Vol. 5, Issue 1
  • DOI: 10.1038/srep15915

Characterization and development of photoactivatable fluorescent proteins for single-molecule-based superresolution imaging
journal, May 2014

  • Wang, S.; Moffitt, J. R.; Dempsey, G. T.
  • Proceedings of the National Academy of Sciences, Vol. 111, Issue 23
  • DOI: 10.1073/pnas.1406593111

Reversible Switching between Nonquenched and Quenched States in Nanoscale Linear Arrays of Plant Light-Harvesting Antenna Complexes
journal, July 2014

  • Vasilev, Cvetelin; Johnson, Matthew P.; Gonzales, Edward
  • Langmuir, Vol. 30, Issue 28
  • DOI: 10.1021/la501483s

Phycobiliproteins — a family of valuable, widely used fluorophores
journal, April 1994

  • Glazer, Alexander N.
  • Journal of Applied Phycology, Vol. 6, Issue 2
  • DOI: 10.1007/BF02186064

Structural Control of the Photodynamics of Boron−Dipyrrin Complexes
journal, November 2005

  • Kee, Hooi Ling; Kirmaier, Christine; Yu, Lianhe
  • The Journal of Physical Chemistry B, Vol. 109, Issue 43
  • DOI: 10.1021/jp0525078

Fluorophore localization algorithms for super-resolution microscopy
journal, February 2014


A far-red fluorescent protein evolved from a cyanobacterial phycobiliprotein
journal, August 2016

  • Rodriguez, Erik A.; Tran, Geraldine N.; Gross, Larry A.
  • Nature Methods, Vol. 13, Issue 9
  • DOI: 10.1038/nmeth.3935

Targeted Green-Red Photoconversion of EosFP, a Fluorescent Marker Protein
journal, December 2005

  • Ivanchenko, Sergey; Röcker, Carlheinz; Oswald, Franz
  • Journal of Biological Physics, Vol. 31, Issue 3-4
  • DOI: 10.1007/s10867-005-0174-z

Small monomeric and highly stable near-infrared fluorescent markers derived from the thermophilic phycobiliprotein, ApcF2
journal, October 2017

  • Ding, Wen-Long; Miao, Dan; Hou, Ya-Nan
  • Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, Vol. 1864, Issue 10
  • DOI: 10.1016/j.bbamcr.2017.08.002

    Figures/Tables have been extracted from DOE-funded journal article accepted manuscripts.