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We present a cryo-EM structure of the monomeric light-harvesting-reaction center (LH1-RC) core complex from photosynthetic purple bacterium Rhodobacter (Rba.) sphaeroides at 2.9 Å resolution. The LH1 complex forms a C-shaped structure composed of 14 αβ-polypeptides around the RC with a large ring opening. From the cryo-EM density map, a previously unrecognized integral membrane protein, referred to as protein-U, was identified. Protein-U has a U-shaped conformation near the LH1-ring opening and was annotated as a hypothetical protein in the Rba. sphaeroides genome. Deletion of protein-U resulted in a mutant strain that expressed a much-reduced amount of the dimeric LH1-RC, indicating an important role for protein-U in dimerization of the LH1-RC complex. PufX was located opposite protein-U on the LH1-ring opening, and both its position and conformation differed from that of previous reports of dimeric LH1-RC structures obtained at low-resolution. Twenty-six molecules of the carotenoid spheroidene arranged in two distinct configurations were resolved in the Rba. sphaeroides LH1 and were positioned within the complex to block its pores. Our findings offer a new view of the core photocomplex of Rba. sphaeroides and the connections between structure and function in bacterial photocomplexes in general.

Rhodospirillum (Rsp.) rubrum is one of the most widely used model organisms in bacterial photosynthesis. This purple phototroph is characterized by the presence of both rhodoquinone (RQ) and ubiquinone as electron carriers and bacteriochlorophyll (BChl) a esterified at the propionic acid side chain by geranylgeraniol (BChl aG) instead of phytol. Despite intensive efforts, the structure of the light-harvesting-reaction center (LH1-RC) core complex from Rsp. rubrum remains at low resolutions. Using cryo-EM, here we present a robust new view of the Rsp. rubrum LH1-RC at 2.76 A resolution. The LH1 complex forms a closed, slightly elliptical ring structure with 16 αβ-polypeptides surrounding the RC. Our biochemical analysis detected RQ molecules in the purified LH1-RC, and the cryo-EM density map specifically positions RQ at the QA site in the RC. The geranylgeraniol side chains of BChl aG coordinated by LH1 β-polypeptides exhibit a highly homologous tail-up conformation that allows for interactions with the bacteriochlorin rings of nearby LH1 α-associated BChls aG. The structure also revealed key protein-protein interactions in both N- and C-terminal regions of the LH1 αβ-polypeptides, mainly within a face-to-face structural subunit. Our high-resolution Rsp. rubrum LH1-RC structure provides new insight for evaluating past experimental and computational results obtained with this old organism over many decades and lays the foundation for more detailed exploration of light-energy conversion, quinone transport, and structure-function relationships in this pigment-protein complex.

Rhodobacter (Rba.) sphaeroides is the most widely used model organism in bacterial photosynthesis. The light-harvesting-reaction center (LH1-RC) core complex of this purple phototroph is characterized by the co-existence of monomeric and dimeric forms, the presence of the protein PufX, and approximately two carotenoids per LH1 αβ-polypeptides. Despite many efforts, structures of the Rba. sphaeroides LH1-RC have not been obtained at high resolutions. Here we report a cryo-EM structure of the monomeric LH1-RC from Rba. sphaeroides strain IL106 at 2.9 Å resolution. The LH1 complex forms a C-shaped structure composed of 14 αβ-polypeptides around the RC with a large ring opening. From the cryo-EM density map, a previously unrecognized integral membrane protein, referred to as protein-U, was identified. Protein-U has a U-shaped conformation near the LH1-ring opening and was annotated as a hypothetical protein in the Rba. sphaeroides genome. Deletion of protein-U resulted in a mutant strain that expressed a much-reduced amount of the dimeric LH1-RC, indicating an important role for protein-U in dimerization of the LH1-RC complex. PufX was located opposite protein-U on the LH1-ring opening, and both its position and conformation differed from that of previous reports of dimeric LH1-RC structures obtained at low-resolution. Twenty-six molecules of the carotenoid spheroidene arranged in two distinct configurations were resolved in the Rba. sphaeroides LH1 and were positioned within the complex to block its channels. Our findings offer an exciting new view of the core photocomplex of Rba. sphaeroides and the connections between structure and function in bacterial photocomplexes in general., Rhodobacter (Rba.) sphaeroides is a model organism for studying bacterial photosynthesis. Here, the authors present the 2.9 Å cryo-EM structure of the monomeric light-harvesting-reaction center core complex from Rba. sphaeroides strain IL106, which revealed the position and conformation of PufX and the presence of an additional component protein-U, an integral membrane protein.

Continuous presentation of the odorant diacetyl to the nematode Caenorhabditis elegans causes a decrease in the level of chemotactic response to diacetyl. This decline in response is caused by diacetyl adaptation. When wild-type nematodes were maintained at 15°C after pre-exposure to diacetyl, diacetyl adaptation did not continue up to 2 hr. Adaptation continued up to 6 hr in nematodes bred at 20°C, and it continued beyond 12 hr in nematodes bred at 25°C. These results indicate that the retention time of diacetyl adaptation is dependent on the environmental breeding temperature and suggest that moderate oxygen signals are required for maintaining the attenuated response to diacetyl because of the correlation between breeding temperature and production of oxygen intermediates. When isp-1 and clk-1 mutants, which show reduced rates of oxygen intermediate production, were maintained at 20 and 25°C after pre-exposure to diacetyl, the mutants showed a shorter retention time of diacetyl adaptation compared with that of wild-type nematodes. When gas-1 and mev-1 mutants, which have a hypersensitive response to oxidative stress, were maintained at 15 and 20°C, they showed a longer retention time of adaptation, that is, adaptation continued beyond 2 and 12 hr, respectively. When wild-type nematodes were maintained on plates that included 0.05% α-lipoic acid, which suppresses production of oxygen intermediates, the retention time of adaptation did not continue up to 6 hr in nematodes bred at 20°C and up to 12 hr in nematodes bred at 25°C. These results support the possibility that oxygen intermediates contribute to retention time for diacetyl adaptation in the nematode C. elegans.

The retention time of attenuated chemotactic response to continuous presentation of odorant diacetyl was investigated in the nematode Caenorhabditis elegans. The level of chemotactic response of nematodes pre-exposed to diacetyl for 90 min was significantly smaller than that of nonexposed control nematodes. In this study, wild-type (N2) nematodes were maintained at 15, 20 and 25 degrees C after pre-exposure to diacetyl. At 20 degrees C, there was a decrease in response to diacetyl continuing for up to 6 hr after pre-exposure to the chemical, but not up to 12 hr. Interestingly, the decrease in response to diacetyl did not continue up to 2 hr in nematodes bred at 15 degrees C, although it continued beyond 12 hr in nematodes bred at 25 degrees C. These results indicate that the retention time of attenuated chemotactic response to diacetyl is dependent on the environmental breeding temperature of nematodes. The breeding temperature correlated with aging speed of nematodes, suggesting that a short life span (higher aging speed) prolongs the retention time of attenuated chemotactic response to diacetyl after pre-exposure to diacetyl. In the long-lived daf-2, age-1, clk-1 and isp-1 mutants, the effect of diacetyl did not continue up to 2 hr. The short-lived daf-16, daf-18, mev-1 and gas-1 mutants showed a longer duration of decrease in response to diacetyl, that is, the retention time of attenuated chemotactic response to diacetyl continued beyond 12 hr. There is a possibility that the duration of decrease in response to diacetyl after pre-exposure to diacetyl was inversely related to the length of nematodes' life span.

SummaryThe chaperonin GroEL interacts with various proteins, leading them to adopt their correct conformations with the aid of GroES and ATP. The actual mechanism is still being debated. In this study, by use of cryo-electron microscopy, we determined the solution structure of the Thermus thermophilus GroEL-GroES complex encapsulating its substrate proteins. We observed the averaged density of substrate proteins in the center of the GroEL-GroES cavity. The position of the averaged substrate density in the cavity suggested a repulsive interaction between a majority of the substrate proteins and the interior wall of the cavity, which is suitable for substrate release. In addition, we observed a distortion of the cis-GroEL ring, especially at the position near the substrate, which indicated that the interaction between the encapsulated proteins and the GroEL ring results in an adjustment in the cavity's shape to accommodate the substrate.