Selective Detection and Assignment of the Solution NMR Signals of Bacteriochlorophyll <i>a</i> in a Reconstituted Subunit of a Light-Harvesting Complex

Wang, Zheng-Yu; Muraoka, Yoshiyuki; Shimonaga, Masahiro; Kobayashi, Masayuki; Nozawa, Tsunenori

doi:10.1021/ja0112994

Cited by 24 publications

(20 citation statements)

References 26 publications

(61 reference statements)

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“…1. The high value of the A 820 /A 270 ratio (2.33), exceeding that of other preparations (33), and the absence of shoulders near 690 nm (chlorin oxidation product) and 770 nm (free BChla) are indicative of the purity of the preparation. The quality of the B820 fraction was confirmed by SDS-PAGE, which showed only two low molecular mass polypeptides at apparent molecular weights between 6 and 10 kDa (not shown).…”

Section: B820mentioning

confidence: 95%

Trapping of an Assembly Intermediate of Photosynthetic LH1 Antenna beyond B820 Subunit

Fiedor

Scheer²

2005

Journal of Biological Chemistry

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Most photosynthetic LH1 antennae undergo dissociation into B820 subunits, suggesting their universal character as structural modules. However, dissociation into subunits seems to occur reversibly only in the absence of carotenoids and the subunits were never found to bind carotenoids. The interactions of carotenoids with B820 have been studied in a newly developed reconstitution assay of the LH1 antenna from Rhodospirillum rubrum (Fiedor, L., Akahane, J., and Koyama, Y. (2004) Biochemistry 43, 16487-16496). These model studies show that B820 subunits strongly interact with carotenoids and spontaneously form stable LH1-like complexes with substoichiometric carotenoid content. This is the first experimental evidence that B820 may occur as a short-lived intermediate in the assembly of LH1 in vivo. The resulting complex of B820 subunits with carotenoid, termed iB873, is homogeneous, according to ion exchange chromatography and reproducible pigment composition. The iB873-bound carotenoid is as efficient in energy transfer to bacteriochlorophyll as the one in native antenna. To our knowledge, iB873 is the first complex binding functional carotenoid, with the spectral and biochemical properties intermediate between that of B820 and the fully assembled LH1.The progress in biochemical (1-3), spectroscopic (4 -8), and structural characterization (9 -16) of purple bacterial lightharvesting complexes has greatly advanced our understanding of photosynthesis. These intramembranous antennae are oligomers of small, heterodimeric proteins carrying BChls 1 and Crts. In peripheral light-harvesting complexes, LH2, LH3, or LH4, 8 or 9 such units assemble to a near perfect circular structure (9 -11). The core light-harvesting complex, LH1, is an oligomer of similar units, but here the size and the shape seem to be more flexible (9 -11,17).To understand the assembly of the pigment-protein complexes, a detailed knowledge is necessary about the role of each particular component in the assembly, and of their interactions in the oligomeric state. The oligomeric character and modularity of antenna structures became already apparent before the high resolution structural data were obtained, mainly because of the isolation of a B820 subunit form of LH1 (2,18,19). Virtually all LH1 complexes were found to undergo dissociation into such subunits, suggesting their function as a universal structural motif (2). They are thought to consist of a dimeric form of BChl, non-covalently attached to two short hydrophobic polypeptides ␣ and ␤, organized into a basic unit ␣␤⅐(BChla) 2 or its dimer (20 -22). B820, however, has mostly been obtained from Crt-less LH1, and only here is the process fully reversible: depending on the detergent concentration, B820 can be reversibly oligomerized in vitro to Crt-less LH1 (low detergent concentration) or dissociated into smaller subunits that still contain monomeric BChl (high detergent concentration) (18,20,21,23). If B820 is prepared by dissociation of Crt-containing LH1, the Crts are lost in the process. In this c...

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Section: B820mentioning

confidence: 95%

Trapping of an Assembly Intermediate of Photosynthetic LH1 Antenna beyond B820 Subunit

Fiedor

Scheer²

2005

Journal of Biological Chemistry

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“…Although we do not precisely know how many OG molecules attach to one B820 subunit, the number of attached OG could be of the same order of magnitude, if not equal, to the aggregation number of micelles formed by OG alone because the LH1 is considered to be a highly hydrophobic integral membrane protein complex. The large molecular weight of B820-containing micelles is also evident from the broad line widths observed in nuclear magnetic resonance spectroscopy (13). The result may explain why there have been large discrepancies in estimation of the size of B820 subunit using the conventional gel exclusion chromatography.…”

Section: Discussionmentioning

confidence: 84%

“…Physical sizes and molecular weights were determined by Guinier analysis without introducing any fitting parameters. Because structural homogeneity is essential for conducting such experiment at high concentrations, we have employed a novel reconstitution method for producing highly homogeneous and concentrated B820 subunits (13), as can be judged from absorption spectra and highresolution nuclear magnetic resonance spectroscopy.…”

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confidence: 99%

Determination of the B820 Subunit Size of a Bacterial Core Light-Harvesting Complex by Small-Angle Neutron Scattering

et al. 2003

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The B820 subunit is an integral pigment-membrane protein complex and can be obtained by both dissociation of the core light-harvesting complex (LH1) in photosynthetic bacteria and reconstitution from its component parts in the presence of n-octyl -D-glucopyranoside (OG). Intrinsic size of the B820 subunit from Rhodospirillum rubrum LH1 complex was measured by small-angle neutron scattering in perdeuterated OG solution and evaluated by Guinier analysis. Both the B820 subunits prepared by dissociation of LH1 and reconstitution from apopolypeptides and pigments were shown to have a molecular weight of 11 400 ( 500 and radius of gyration of 11.0 ( 1.0 Å, corresponding to a heterodimer consisting of one pair of R -polypeptides and two bacteriochlorophyll a molecules. Molecular weights of micelles formed by OG alone in solutions were determined in a range from 30 000 to 50 000 over concentrations of 1-5% (w/v), and thus are much larger than that of the B820 subunit. Similar measurement on the pigment-depleted apopolypeptides revealed highly heterogeneous behavior in the OG solutions, indicating that aggregates with various sizes were formed. The result provides evidence that bacteriochlorophyll a molecules play a crucial role in stabilizing and maintaining the B820 subunits in the dimeric state in solution. Further measurements on individual R-and -polypeptides exhibited a marked difference in aggregation property between the two polypeptides. The R-polypeptides appear to be uniformly dissolved in OG solution in a monomeric form, whereas the -polypeptides favor a self-associated form and tend to form large aggregates even in the presence of detergent. The difference in aggregation tendency was discussed in relation to the different behavior between R-and -polypeptides in reconstitution with bacteriochlorophyll a molecules.Despite intense interest and continuing investigation, controversy over the nature of an intermediate form of bacterial core light-harvesting complexes (LH1 1 ) has not been settled. The LH1 complex, serving as highly efficient molecular machinery for collection and transfer of solar energy to photochemical reaction centers in photosynthetic bacteria, is a large multimer of a minimal unit composed of two small integral membrane polypeptides (R and , ca. 6 kDa) associated with two bacteriochlorophyll (BChl) and one carotenoid molecule. Upon addition of detergents, the LH1 multimer can be dissociated into a structural subunit characterized by a Q y absorption band at 820 nm (1). Both the subunit (referred to as B820) and LH1 complex can be reversibly dissociated and reconstituted from the their components (2). Early chromatographic studies by gel filtration estimated a range of molecular weight from 32 to 65 kDa for the B820 subunit, roughly corresponding to a tetramer composed of two minimal structural units (1, 3). In contrast, spectroscopic evidence suggested that the B820 is a heterodimer consisting of two interacting BChl a molecules linked to one R -polypeptide pair (4,5). Recently, ide...

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“…Due to their less-ordered oligomer structures, crystallization of the LH1 protein complexes has only resulted in low-resolution structures (Roszak et al 2003; Karrasch et al 1995). The LH1 structural inhomogeneity similarly induces broadening of the lines in the NMR spectra and in an earlier study on Rhodospirillum rubrum LH1 αβ subunits reconstituted with 13 C– 15 N-labeled BChls, only one set of BChl NMR signals was assigned without distinction between the α - and β -BChl (Wang et al 2002). In our recent work, NMR assignment of the two types of LH1 BChls was achieved in intact LH1-RC core complexes, using the LH2 spectra as a “template” for the assignment.…”

Section: Introductionmentioning

confidence: 94%

Solid-state NMR applied to photosynthetic light-harvesting complexes

Pandit

Groot

2011

Photosynth Res

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This short review describes how solid-state NMR has provided a mechanistic and electronic picture of pigment–protein and pigment–pigment interactions in photosynthetic antenna complexes. NMR results on purple bacterial antenna complexes show how the packing of the protein and the pigments inside the light-harvesting oligomers induces mutual conformational stress. The protein scaffold produces deformation and electrostatic polarization of the BChl macrocycles and leads to a partial electronic charge transfer between the BChls and their coordinating histidines, which can tune the light-harvesting function. In chlorosome antennae assemblies, the NMR template structure reveals how the chromophores can direct their self-assembly into higher macrostructures which, in turn, tune the light-harvesting properties of the individual molecules by controlling their disorder, structural deformation, and electronic polarization without the need for a protein scaffold. These results pave the way for addressing the next challenge, which is to resolve the functional conformational dynamics of the lhc antennae of oxygenic species that allows them to switch between light-emitting and light-energy dissipating states.

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Selective Detection and Assignment of the Solution NMR Signals of Bacteriochlorophyll a in a Reconstituted Subunit of a Light-Harvesting Complex

Cited by 24 publications

References 26 publications

Trapping of an Assembly Intermediate of Photosynthetic LH1 Antenna beyond B820 Subunit

Trapping of an Assembly Intermediate of Photosynthetic LH1 Antenna beyond B820 Subunit

Determination of the B820 Subunit Size of a Bacterial Core Light-Harvesting Complex by Small-Angle Neutron Scattering

Solid-state NMR applied to photosynthetic light-harvesting complexes

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