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Title: Single-molecule trapping and spectroscopy reveals photophysical heterogeneity of phycobilisomes quenched by Orange Carotenoid Protein

Abstract

The Orange Carotenoid Protein (OCP) is a cytosolic photosensor that is responsible for nonphotochemical quenching (NPQ) of the light-harvesting process in most cyanobacteria. Upon photoactivation by blue-green light, OCP binds to the phycobilisome antenna complex, providing an excitonic trap to thermally dissipate excess energy. At present, both the binding site and NPQ mechanism of OCP are unknown. Using an Anti-Brownian ELectrokinetic (ABEL) trap, we isolate single phycobilisomes in free solution, both in the presence and absence of activated OCP, to directly determine the photophysics and heterogeneity of OCPquenched phycobilisomes. Surprisingly, we observe two distinct OCP-quenched states, with lifetimes 0.09 ns (6% of unquenched brightness) and 0.21 ns (11% brightness). Photon-byphoton Monte Carlo simulations of exciton transfer through the phycobilisome suggest that the observed quenched states are kinetically consistent with either two or one bound OCPs, respectively, underscoring an additional mechanism for excitation control in this key photosynthetic unit.

Authors:
ORCiD logo [1];  [1]; ORCiD logo [2];  [2]; ORCiD logo [2]; ORCiD logo [1]
  1. Stanford Univ., CA (United States). Dept. of Chemistry
  2. Washington Univ., St. Louis, MO (United States). Dept. of Biology and Chemistry
Publication Date:
Research Org.:
Stanford Univ., CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC)
OSTI Identifier:
1501915
Grant/Contract Number:  
FG02-07ER15892
Resource Type:
Accepted Manuscript
Journal Name:
Nature Communications
Additional Journal Information:
Journal Volume: 10; Journal Issue: 1; Journal ID: ISSN 2041-1723
Publisher:
Nature Publishing Group
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Squires, Allison H., Dahlberg, Peter D., Liu, Haijun, Magdaong, Nikki Cecil M., Blankenship, Robert E., and Moerner, W. E. Single-molecule trapping and spectroscopy reveals photophysical heterogeneity of phycobilisomes quenched by Orange Carotenoid Protein. United States: N. p., 2019. Web. doi:10.1038/s41467-019-09084-2.
Squires, Allison H., Dahlberg, Peter D., Liu, Haijun, Magdaong, Nikki Cecil M., Blankenship, Robert E., & Moerner, W. E. Single-molecule trapping and spectroscopy reveals photophysical heterogeneity of phycobilisomes quenched by Orange Carotenoid Protein. United States. doi:10.1038/s41467-019-09084-2.
Squires, Allison H., Dahlberg, Peter D., Liu, Haijun, Magdaong, Nikki Cecil M., Blankenship, Robert E., and Moerner, W. E. Tue . "Single-molecule trapping and spectroscopy reveals photophysical heterogeneity of phycobilisomes quenched by Orange Carotenoid Protein". United States. doi:10.1038/s41467-019-09084-2. https://www.osti.gov/servlets/purl/1501915.
@article{osti_1501915,
title = {Single-molecule trapping and spectroscopy reveals photophysical heterogeneity of phycobilisomes quenched by Orange Carotenoid Protein},
author = {Squires, Allison H. and Dahlberg, Peter D. and Liu, Haijun and Magdaong, Nikki Cecil M. and Blankenship, Robert E. and Moerner, W. E.},
abstractNote = {The Orange Carotenoid Protein (OCP) is a cytosolic photosensor that is responsible for nonphotochemical quenching (NPQ) of the light-harvesting process in most cyanobacteria. Upon photoactivation by blue-green light, OCP binds to the phycobilisome antenna complex, providing an excitonic trap to thermally dissipate excess energy. At present, both the binding site and NPQ mechanism of OCP are unknown. Using an Anti-Brownian ELectrokinetic (ABEL) trap, we isolate single phycobilisomes in free solution, both in the presence and absence of activated OCP, to directly determine the photophysics and heterogeneity of OCPquenched phycobilisomes. Surprisingly, we observe two distinct OCP-quenched states, with lifetimes 0.09 ns (6% of unquenched brightness) and 0.21 ns (11% brightness). Photon-byphoton Monte Carlo simulations of exciton transfer through the phycobilisome suggest that the observed quenched states are kinetically consistent with either two or one bound OCPs, respectively, underscoring an additional mechanism for excitation control in this key photosynthetic unit.},
doi = {10.1038/s41467-019-09084-2},
journal = {Nature Communications},
number = 1,
volume = 10,
place = {United States},
year = {2019},
month = {3}
}

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Figures / Tables:

Fig. 1 Fig. 1 : OCP quenches fluorescence in the phycobilisome. a Structure of OCP in inactive orange form, from PDB 3MG142. A keto-carotenoid spans the interior of the N- and C-terminal domains (NTD and CTD, respectively), which are connected with a flexible linker and separate upon photoactivation. b Schematic of OCPmore » activation. Upon absorption of blue-green light, OCPO is converted to OCPR, which binds to and quenches an unknown location on the core of the phycobilisome (PB). c Structure of the truncated PB mutant used in this study (CB-PB from Synechocystis PCC 680351). Disc-shaped trimers of phycobiliproteins are joined by linker proteins into stacks that make up the core and rods of the PB. d Bulk fluorescence emission spectrum of PB in the absence (navy) and presence (maroon) of bound OCPR. Solid lines: area-normalized for total relative fluorescence intensity. Dotted line: peak-normalized. e Bulk lifetime decays for PB in the absence (light blue) and presence (light red) of bound OCPR. Single- and double-exponential fits are shown in navy and maroon, respectively. Gray: instrument response function, FWHM 50 ps« less

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