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Title: Direct single-molecule measurements of phycocyanobilin photophysics in monomeric C-phycocyanin

Abstract

Phycobilisomes are highly organized pigment–protein antenna complexes found in the photosynthetic apparatus of cyanobacteria and rhodophyta that harvest solar energy and transport it to the reaction center. A detailed bottom-up model of pigment organization and energy transfer in phycobilisomes is essential to understanding photosynthesis in these organisms and informing rational design of artificial light-harvesting systems. In particular, heterogeneous photophysical behaviors of these proteins, which cannot be predicted de novo, may play an essential role in rapid light adaptation and photoprotection. Furthermore, the delicate architecture of these pigment–protein scaffolds sensitizes them to external perturbations, for example, surface attachment, which can be avoided by study in free solution or in vivo. Here, we present single-molecule characterization of C-phycocyanin (C-PC), a three-pigment biliprotein that self-assembles to form the midantenna rods of cyanobacterial phycobilisomes. Using the Anti-Brownian Electrokinetic (ABEL) trap to counteract Brownian motion of single particles in real time, we directly monitor the changing photophysical states of individual C-PC monomers from Spirulina platensis in free solution by simultaneous readout of their brightness, fluorescence anisotropy, fluorescence lifetime, and emission spectra. These include single-chromophore emission states for each of the three covalently bound phycocyanobilins, providing direct measurements of the spectra and photophysics of these chemicallymore » identical molecules in their native protein environment. We further show that a simple Förster resonant energy transfer (FRET) network model accurately predicts the observed photophysical states of C-PC and suggests highly variable quenching behavior of one of the chromophores, which should inform future studies of higher-order complexes.« less

Authors:
;
Publication Date:
Research Org.:
Stanford Univ., CA (United States). Dept. of Chemistry
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22)
OSTI Identifier:
1377085
Alternate Identifier(s):
OSTI ID: 1377406
Grant/Contract Number:  
FG02-07ER15892
Resource Type:
Published Article
Journal Name:
Proceedings of the National Academy of Sciences of the United States of America
Additional Journal Information:
Journal Name: Proceedings of the National Academy of Sciences of the United States of America Journal Volume: 114 Journal Issue: 37; Journal ID: ISSN 0027-8424
Publisher:
Proceedings of the National Academy of Sciences
Country of Publication:
United States
Language:
English
Subject:
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY; 59 BASIC BIOLOGICAL SCIENCES; photosynthetic protein; C-phycocyanin; ABEL trap; single-molecule spectroscopy; photodynamics

Citation Formats

Squires, Allison H., and Moerner, W. E. Direct single-molecule measurements of phycocyanobilin photophysics in monomeric C-phycocyanin. United States: N. p., 2017. Web. doi:10.1073/pnas.1705435114.
Squires, Allison H., & Moerner, W. E. Direct single-molecule measurements of phycocyanobilin photophysics in monomeric C-phycocyanin. United States. doi:10.1073/pnas.1705435114.
Squires, Allison H., and Moerner, W. E. Mon . "Direct single-molecule measurements of phycocyanobilin photophysics in monomeric C-phycocyanin". United States. doi:10.1073/pnas.1705435114.
@article{osti_1377085,
title = {Direct single-molecule measurements of phycocyanobilin photophysics in monomeric C-phycocyanin},
author = {Squires, Allison H. and Moerner, W. E.},
abstractNote = {Phycobilisomes are highly organized pigment–protein antenna complexes found in the photosynthetic apparatus of cyanobacteria and rhodophyta that harvest solar energy and transport it to the reaction center. A detailed bottom-up model of pigment organization and energy transfer in phycobilisomes is essential to understanding photosynthesis in these organisms and informing rational design of artificial light-harvesting systems. In particular, heterogeneous photophysical behaviors of these proteins, which cannot be predicted de novo, may play an essential role in rapid light adaptation and photoprotection. Furthermore, the delicate architecture of these pigment–protein scaffolds sensitizes them to external perturbations, for example, surface attachment, which can be avoided by study in free solution or in vivo. Here, we present single-molecule characterization of C-phycocyanin (C-PC), a three-pigment biliprotein that self-assembles to form the midantenna rods of cyanobacterial phycobilisomes. Using the Anti-Brownian Electrokinetic (ABEL) trap to counteract Brownian motion of single particles in real time, we directly monitor the changing photophysical states of individual C-PC monomers from Spirulina platensis in free solution by simultaneous readout of their brightness, fluorescence anisotropy, fluorescence lifetime, and emission spectra. These include single-chromophore emission states for each of the three covalently bound phycocyanobilins, providing direct measurements of the spectra and photophysics of these chemically identical molecules in their native protein environment. We further show that a simple Förster resonant energy transfer (FRET) network model accurately predicts the observed photophysical states of C-PC and suggests highly variable quenching behavior of one of the chromophores, which should inform future studies of higher-order complexes.},
doi = {10.1073/pnas.1705435114},
journal = {Proceedings of the National Academy of Sciences of the United States of America},
number = 37,
volume = 114,
place = {United States},
year = {2017},
month = {8}
}

Journal Article:
Free Publicly Available Full Text
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DOI: 10.1073/pnas.1705435114

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