Calcium ions play a dual role in expanding the spectral diversity and structural stability of photocomplexes from several Ca2+-requiring purple sulfur phototrophic bacteria. Here, metal-sensitive structural changes in the isotopically labeled light-harvesting 1 reaction center (LH1-RC) complexes from the thermophilic purple sulfur bacterium Thermochromatium (Tch.) tepidum were investigated by perfusion-induced attenuated total reflection (ATR) Fourier transform infrared (FTIR) spectroscopy. The ATR-FTIR difference spectra induced by exchanges between native Ca2+ and exogenous Ba2+ exhibited interconvertible structural and/or conformational changes in the metal binding sites at the LH1 C-terminal region. Most of the characteristic Ba2+/Ca2+ difference bands were detected even when only Ca ions were removed from the LH1-RC complexes, strongly indicating the pivotal roles of Ca2+ in maintaining the LH1-RC structure of Tch. tepidum. Upon 15N-, 13C- or 2H-labeling, the LH1-RC complexes exhibited characteristic 15N/14N-, 13C/12C-, or 2H/1H-isotopic shifts for the Ba2+/Ca2+ difference bands. Some of the 15N/14N or 13C/12C bands were also sensitive to further 2H-labelings. Given the band frequencies and their isotopic shifts along with the structural information of the Tch. tepidum LH1-RC complexes, metal-sensitive FTIR bands were tentatively identified to the vibrational modes of the polypeptide main chains and side chains comprising the metal binding sites. Furthermore, important new IR marker bands highly sensitive to the LH1 BChl a conformation in the Ca2+-bound states were revealed based on both ATR-FTIR and near-infrared Raman analyses. The present approach provides valuable insights concerning the dynamic equilibrium between the Ca2+- and Ba2+-bound states statically resolved by x-ray crystallography.
REFERENCES
1.
S.
Niwa
, L.-J.
Yu
, K.
Takeda
, Y.
Hirano
, T.
Kawakami
, Z.-Y.
Wang-Otomo
, and K.
Miki
, “Structure of the LH1–RC complex from Thermochromatium tepidum at 3.0 Å
,” Nature
508
, 228
–232
(2014
).2.
L.-J.
Yu
, M.
Suga
, Z.-Y.
Wang-Otomo
, and J.-R.
Shen
, “Structure of photosynthetic LH1–RC super-complex at 1.9 Å resolution
,” Nature
556
, 209
–213
(2018
).3.
P.
Qian
, M. Z.
Papiz
, P. J.
Jackson
, A. A.
Brindley
, I. W.
Ng
, J. D.
Olsen
, M. J.
Dickman
, P. A.
Bullough
, and C. N.
Hunter
, “Three-dimensional structure of the Rhodobacter sphaeroides RC-LH1-PufX complex: Dimerization and quinone channels promoted by PufX
,” Biochemistry
52
, 7575
–7585
(2013
).4.
Y.
Xin
, Y.
Shi
, T.
Niu
, Q.
Wang
, W.
Niu
, X.
Huang
, W.
Ding
, L.
Yang
, R. E.
Blankenship
, X.
Xu
, and F.
Sun
, “Cryo-EM structure of the RC-LH core complex from an early branching photosynthetic prokaryote
,” Nat. Commun.
9
, 1568
(2018
).5.
K.
Tani
, R.
Kanno
, Y.
Makino
, M.
Hall
, M.
Takenouchi
, M.
Imanishi
, L.-J.
Yu
, J.
Overmann
, M. T.
Madigan
, Y.
Kimura
, A.
Mizoguchi
, B. M.
Humbel
, and Z.-Y.
Wang-Otomo
, “Cryo-EM structure of a Ca2+-bound photosynthetic LH1-RC complex containing multiple αβ-polypeptides
,” Nat. Commun.
11
, 4955
(2020
).6.
K.
Tani
, R.
Kanno
, X.-C.
Ji
, M.
Hall
, L.-J.
Yu
, Y.
Kimura
, M. T.
Madigan
, A.
Mizoguchi
, B. M.
Humbel
, and Z.-Y.
Wang-Otomo
, “Cryo-EM structure of the photosynthetic LH1-RC complex from Rhodospirillum rubrum
,” Biochemistry
60
, 2483
–2491
(2021
).7.
P.
Qian
, T. I.
Croll
, D. J. K.
Swainsbury
, P.
Castro-Hartmann
, N. W.
Moriarty
, K.
Sader
, and C. N.
Hunter
, “Cryo-EM structure of the Rhodospirillum rubrum RC-LH1 complex at 2.5 Å
,” Biochem. J.
478
, 3253
–3263
(2021
).8.
L.
Bracun
, A.
Yamagata
, B. M.
Christianson
, T.
Terada
, D. P.
Canniffe
, M.
Shirouzu
, and L. N.
Liu
, “Cryo-EM structure of the photosynthetic RC-LH1-PufX supercomplex at 2.8-Å resolution
,” Sci. Adv.
7
, eabf8864
(2021
).9.
D. J. K.
Swainsbury
, P.
Qian
, P. J.
Jackson
, K. M.
Faries
, D. M.
Niedzwiedzki
, E. C.
Martin
, D. A.
Farmer
, L. A.
Malone
, R. F.
Thompson
, N. A.
Ranson
, D. P.
Canniffe
, M. J.
Dickman
, D.
Holten
, C.
Kirmaier
, A.
Hitchcock
, and C. N.
Hunter
, “Structures of Rhodopseudomonas palustris RC-LH1 complexes with open or closed quinone channels
,” Sci. Adv.
7
, eabe2631
(2021
).10.
A.
Hiraishi
, “Transfer of the bacteriochlorophyll b-containing phototrophic bacteria Rhodopseudomonas viridis and Rhodopseudomonas sulfoviridis to the genus Blastochloris gen. nov.
,” Int. J. Syst. Bacteriol.
47
, 217
–219
(1997
).11.
S. M.
Resnick
and M. T.
Madigan
, “Isolation and characterization of a mildly thermophilic nonsulfur purple bacterium containing bacteriochlorophyll b
,” FEMS Microbiol. Lett.
65
, 165
–170
(1989
).12.
J. F.
Imhoff
and H. G.
Trüper
, “Ectothiorhodospira abdelmalekii sp. nov., a new halophilic and alkaliphilic phototropic bacterium
,” Zentralbl. Bakteriol. Mikrobiol. Hyg.
2
, 228
–234
(1981
).13.
J. F.
Imhoff
and H. G.
Trüper
, “Ectothiorhodospira halochloris sp. nov., a new extremely halophilic phototropic bacterium containing bacteriochlorophyll b
,” Arch. Microbiol.
114
, 115
–121
(1977
).14.
C.
Tuschak
, J. T.
Beatty
, and J.
Overmann
, “Photosynthesis genes and LH1 proteins of Roseospirillum parvum 930I, a purple non-sulfur bacterium with unusual spectral properties
,” Photosynth. Res.
81
, 181
–199
(2004
).15.
M. T.
Madigan
, “A novel photosynthetic purple bacterium isolated from a Yellowstone hot spring
,” Science
225
, 313
–315
(1984
).16.
M. T.
Madigan
, “Anoxygenic phototrophic bacteria from extreme environments
,” Photosynth. Res.
76
, 157
–171
(2003
).17.
H. P.
Permentier
, S.
Neerken
, J.
Overmann
, and J.
Amesz
, “A bacteriochlorophyll a antenna complex from purple bacteria absorbing at 963 nm
,” Biochemistry
40
, 5573
–5578
(2001
).18.
O.
Rücker
, A.
Köhler
, B.
Behammer
, K.
Sichau
, and J.
Overmann
, “Puf operon sequences and inferred structures of light-harvesting complexes of three closely related Chromatiaceae exhibiting different absorption characteristics
,” Arch. Microbiol.
194
, 123
–134
(2012
).19.
Y.
Kimura
, Y.
Hirano
, L.-J.
Yu
, H.
Suzuki
, M.
Kobayashi
, and Z.-Y.
Wang
, “Calcium ions are involved in the unusual red shift of the light-harvesting 1 Qy transition of the core complex in thermophilic purple sulfur bacterium Thermochromatium tepidum
,” J. Biol. Chem.
283
, 13867
–13873
(2008
).20.
M.
Imanishi
, M.
Takenouchi
, S.
Takaichi
, S.
Nakagawa
, Y.
Saga
, S.
Takenaka
, M. T.
Madigan
, J.
Overmann
, Z.-Y.
Wang-Otomo
, and Y.
Kimura
, “A dual role for Ca2+ in expanding the spectral diversity and stability of light-harvesting 1 reaction center photocomplexes of purple phototrophic bacteria
,” Biochemistry
58
, 2844
–2852
(2019
).21.
R.
Seto
, S.
Takaichi
, T.
Kurihara
, R.
Kishi
, M.
Honda
, S.
Takenaka
, Y.
Tsukatani
, M. T.
Madigan
, Z.-Y.
Wang-Otomo
, and Y.
Kimura
, “Lycopene-family carotenoids confer thermostability on photocomplexes from a new thermophilic purple bacterium
,” Biochemistry
59
, 2351
–2358
(2020
).22.
M. T.
Madigan
, J. N.
Crespi
, J. E.
Mayers
, M.
Asao
, D. O.
Jung
, K. S.
Bender
, M. L.
Kempher
, M. K.
Hayward
, S. A.
Sanguedolce
, A. C.
Brown
, S.
Takaichi
, Y.
Tsukatani
, H.
Mori
, Z.-Y.
Wang-Otomo
, D. M.
Ward
, and W. M.
Sattley
, “Allochromatium tepidum, sp. nov., a new species of thermophilic purple sulfur bacteria
,” Arch. Microbiol.
204
, 115
(2022
).23.
Y.
Kimura
, L.-J.
Yu
, Y.
Hirano
, H.
Suzuki
, and Z.-Y.
Wang
, “Calcium ions are required for the enhanced thermal stability of the light-harvesting-reaction center core complex from thermophilic purple sulfur bacterium Thermochromatium tepidum
,” J. Biol. Chem.
284
, 93
–99
(2009
).24.
Y.
Kimura
, S.
Lyu
, A.
Okoshi
, K.
Okazaki
, N.
Nakamura
, A.
Ohashi
, T.
Ohno
, M.
Kobayashi
, M.
Imanishi
, S.
Takaichi
, M. T.
Madigan
, and Z.-Y.
Wang-Otomo
, “Effects of calcium ions on the thermostability and spectroscopic properties of the LH1-RC complex from a new thermophilic purple bacterium Allochromatium tepidum
,” J. Phys. Chem. B
121
, 5025
–5032
(2017
).25.
L.-J.
Yu
, T.
Kawakami
, Y.
Kimura
, and Z.-Y.
Wang-Otomo
, “Structural basis for the unusual Qy red-shift and enhanced thermostability of the LH1 complex from Thermochromatium tepidum
,” Biochemistry
55
, 6495
–6504
(2016
).26.
A.
Barth
, “The infrared absorption of amino acid side chains
,” Prog. Biophys. Mol. Biol.
74
, 141
–173
(2000
).27.
A.
Barth
and C.
Zscherp
, “What vibrations tell us about proteins
,” Q. Rev. Biophys.
35
, 369
–430
(2002
).28.
R.
Debus
, “Protein ligation of the photosynthetic oxygen-evolving center
,” Coord. Chem. Rev.
252
, 244
–258
(2008
).29.
T.
Noguchi
, “Fourier transform infrared analysis of the photosynthetic oxygen-evolving center
,” Coord. Chem. Rev.
252
, 336
–346
(2008
).30.
A.
Mezzetti
, “Light-induced infrared difference spectroscopy in the investigation of light harvesting complexes
,” Molecules
20
, 12229
–12249
(2015
).31.
H.
Kandori
, “Structure/function study of photoreceptive proteins by FTIR spectroscopy
,” Bull. Chem. Soc. Jpn.
93
, 904
–926
(2020
).32.
J. E.
Baenziger
, K. W.
Miller
, M. P.
Mccarthy
, and K. J.
Rothschild
, “Probing conformational-changes in the nicotinic acetylcholine-receptor by Fourier-transform infrared difference spectroscopy
,” Biophys. J.
62
, 64
–66
(1992
).33.
R. M.
Nyquist
, D.
Heitbrink
, C.
Bolwien
, T. A.
Wells
, R. B.
Gennis
, and J.
Heberle
, “Perfusion-induced redox differences in cytochrome c oxidase: ATR/FT-IR spectroscopy
,” FEBS Lett.
505
, 63
–67
(2001
).34.
M.
Iwaki
, J.
Breton
, and P. R.
Rich
, “ATR-FTIR difference spectroscopy of the PM intermediate of bovine cytochrome c oxidase
,” Biochim. Biophys. Acta, Bioenerg.
1555
, 116
–121
(2002
).35.
S. E.
Ryan
, D. G.
Hill
, and J. E.
Baenziger
, “Dissecting the chemistry of nicotinic receptor-ligand interactions with infrared difference spectroscopy
,” J. Biol. Chem.
277
, 10420
–10426
(2002
).36.
R. M.
Nyquist
, D.
Heitbrink
, C.
Bolwien
, R. B.
Gennis
, and J.
Heberle
, “Direct observation of protonation reactions during the catalytic cycle of cytochrome c oxidase
,” Proc. Natl. Acad. Sci. U. S. A.
100
, 8715
–8720
(2003
).37.
M.
Iwaki
and P. R.
Rich
, “Direct detection of formate ligation in cytochrome c oxidase by ATR-FTIR spectroscopy
,” J. Am. Chem. Soc.
126
, 2386
–2389
(2004
).38.
J.
Guijarro
, M.
Engelhard
, and F.
Siebert
, “Anion uptake in halorhodopsin from Natromonas pharaonis studied by FTIR spectroscopy: Consequences for the anion transport mechanism
,” Biochemistry
45
, 11578
–11588
(2006
).39.
M.
Iwaki
, N. P. J.
Cotton
, P. G.
Quirk
, P. R.
Rich
, and J. B.
Jackson
, “Molecular recognition between protein and nicotinamide dinucleotide in intact, proton-translocating transhydrogenase studied by ATR-FTIR spectroscopy
,” J. Am. Chem. Soc.
128
, 2621
–2629
(2006
).40.
Y.
Furutani
, T.
Murata
, and H.
Kandori
, “Sodium or lithium ion-binding-induced structural changes in the K-ring of V-ATPase from Enterococcus hirae revealed by ATR-FTIR spectroscopy
,” J. Am. Chem. Soc.
133
, 2860
–2863
(2011
).41.
A.
Maréchal
, M.
Iwaki
, and P. R.
Rich
, “Structural changes in cytochrome c oxidase induced by binding of sodium and calcium ions: An ATR-FTIR study
,” J. Am. Chem. Soc.
135
, 5802
–5807
(2013
).42.
M.
Iwaki
, G.
Yakovlev
, J.
Hirst
, A.
Osyczka
, P. L.
Dutton
, D.
Marshall
, and P. R.
Rich
, “Direct observation of redox-linked histidine protonation changes in the iron–sulfur protein of the cytochrome bc1 complex by ATR-FTIR spectroscopy
,” Biochemistry
44
, 4230
–4237
(2005
).43.
M.
Iwaki
, S.
Andrianambinintsoa
, P.
Rich
, and J.
Breton
, “Attenuated total reflection Fourier transform infrared spectroscopy of redox transitions in photosynthetic reaction centers: Comparison of perfusion- and light-induced difference spectra
,” Spectrochim. Acta, Part A
58
, 1523
–1533
(2002
).44.
I.
Oda
, M.
Iwaki
, D.
Fujita
, Y.
Tsutsui
, S.
Ishizaka
, M.
Dewa
, M.
Nango
, T.
Kajino
, Y.
Fukushima
, and S.
Itoh
, “Photosynthetic electron transfer from reaction center pigment-protein complex in silica nanopores
,” Langmuir
26
, 13399
–13406
(2010
).45.
Y.
Li
, Y.
Kimura
, T.
Arikawa
, Z.-Y.
Wang-Otomo
, and T.
Ohno
, “ATR–FTIR detection of metal-sensitive structural changes in the light-harvesting 1 reaction center complex from the thermophilic purple sulfur bacterium Thermochromatium tepidum
,” Biochemistry
52
, 9001
–9008
(2013
).46.
Y.
Kimura
, Y.
Yura
, Y.
Hayashi
, Y.
Li
, M.
Onoda
, L.-J.
Yu
, Z.-Y.
Wang-Otomo
, and T.
Ohno
, “Spectroscopic and thermodynamic characterization of the metal-binding sites in the LH1–RC complex from thermophilic photosynthetic bacterium Thermochromatium tepidum
,” J. Phys. Chem. B
120
, 12466
–12473
(2016
).47.
T.
Noguchi
and M.
Sugiura
, “Analysis of flash-induced FTIR difference spectra of the S-state cycle in the photosynthetic water-oxidizing complex by uniform 15N- and 13C-isotope labeling
,” Biochemistry
42
, 6035
–6042
(2003
).48.
Y.
Kimura
, N.
Mizusawa
, A.
Ishii
, T.
Yamanari
, and T.-a.
Ono
, “Changes of low-frequency vibrational modes induced by universal 15N- and 13C-isotope labeling in S2/S1 FTIR difference spectrum of oxygen-evolving complex
,” Biochemistry
42
, 13170
–13177
(2003
).49.
B. B.
Ivanova
, “IR-LD spectroscopic characterization of L-Tryptophan containing dipeptides
,” Spectrochim. Acta, Part A
64
, 931
–938
(2006
).50.
B.
Hernández
, F.
Pflüger
, A.
Adenier
, S. G.
Kruglik
, and M.
Ghomi
, “Vibrational analysis of amino acids and short peptides in hydrated media. VIII. Amino acids with aromatic side chains: L-phenylalanine, L-tyrosine, and L-tryptophan
,” J. Phys. Chem. B
114
, 15319
–15330
(2010
).51.
G.
Socrates
, Infrared and Raman Characteristic Group Frequencies: Tables and Charts
, 3rd ed. (Wiley
, Chichester; New York
, 2001
).52.
G.
Deacon
and R. J.
Phillips
, “Relationships between the carbon-oxygen stretching frequencies of carboxylato complexes and the type of carboxylate coordination
,” Coord. Chem. Rev.
33
, 227
–250
(1980
).53.
K.
Nakamoto
, Infrared and Raman Spectra of Inorganic and Coordination Compounds, Part B: Applications in Coordination, Organometallic, and Bioinorganic Chemistry
, 5th ed. (John Wiley & Sons
, New York
, 1997
), pp. 59
–62
.54.
W. G.
Mantele
, A. M.
Wollenweber
, E.
Nabedryk
, and J.
Breton
, “Infrared spectroelectrochemistry of bacteriochlorophylls and bacteriopheophytins: Implications for the binding of the pigments in the reaction center from photosynthetic bacteria
,” Proc. Natl. Acad. Sci. U. S. A.
85
, 8468
–8472
(1988
).55.
T. A.
Mattioli
, A.
Hoffmann
, B.
Robert
, B.
Schrader
, and M.
Lutz
, “Primary donor structure and interactions in bacterial reaction centers from near-infrared Fourier-transform resonance Raman-spectroscopy
,” Biochemistry
30
, 4648
–4654
(1991
).56.
T.
Noguchi
, Y.
Furukawa
, and M.
Tasumi
, “1064-nm-excited Fourier-transform Raman studies of bacteriochlorophyll-a in solid films and in a blue-green mutant of Rhodobacter sphaeroides
,” Spectrochim. Acta, Part A
47
, 1431
–1440
(1991
).57.
M.
Ceccarelli
, M.
Lutz
, and M.
Marchi
, “A density functional normal mode calculation of a bacteriochlorophyll a derivative
,” J. Am. Chem. Soc.
122
, 3532
–3533
(2000
).58.
F.
Wei
, D.
Zhang
, N. J.
Halas
, and J. D.
Hartgerink
, “Aromatic amino acids providing characteristic motifs in the Raman and SERS spectroscopy of peptides
,” J. Phys. Chem. B
112
, 9158
–9164
(2008
).59.
M.
Wolpert
and P.
Hellwig
, “Infrared spectra and molar absorption coefficients of the 20 alpha amino acids in aqueous solutions in the spectral range from 1800 to 500 cm−1
,” Spectrochim. Acta, Part A
64
, 987
–1001
(2006
).© 2022 Author(s). Published under an exclusive license by AIP Publishing.
2022
Author(s)
You do not currently have access to this content.