SummaryPhotosystem II (PSII) is a huge membrane-protein complex consisting of 20 different subunits with a total molecular mass of 350 kDa for a monomer, and catalyzes light-driven water oxidation at its catalytic center, the oxygen-evolving complex (OEC) [1][2][3] . The structure of PSII has been analyzed at 1.9 Å resolution by synchrotron radiation X-rays, which revealed that OEC is a Mn4CaO5 cluster organized in an asymmetric, "distorted-chair" form 4 . This structure was further analyzed with femtosecond X-ray free electron lasers (XFEL), providing the "radiation damage-free" 5 structure. The mechanism of O=O bond formation, however, remains obscure due to the lack of intermediate state structures. Here we report the structural changes of PSII induced by 2-flash (2F) illumination at room temperature at a resolution of 2.35 Å using time-resolved serial femtosecond crystallography (TR-SFX) with an XFEL provided by the SPring-8 angstrom compact free-electron laser (SACLA). Isomorphous differenceFourier map between the 2F and dark-adapted states revealed two areas of apparent changes; they are around QB/non-heme iron and the Mn4CaO5 cluster. The changes around the QB/non-heme iron region reflected the electron and proton transfers induced by the 2F-illumination. In the region around the Mn4CaO5 cluster, a water molecule located 3.5 Å from the Mn4CaO5 cluster disappeared from the map upon 2Fillumination, leading to a closer distance between another water molecule and O4, suggesting also the occurrence of proton transfer. Importantly, the 2F-dark isomorphous difference Fourier map showed an apparent positive peak around O5, a unique μ3-oxo-bridge located in the quasi-center of Mn1 and Mn4 4,5 . This suggests an insertion of a new oxygen atom (O6) close to O5, providing an O=O distance of 1.5 Å between these two oxygen atoms. This provides a mechanism for the O=O bond formation 4 consistent with that proposed by Siegbahn 6,7 . Fig. 1a shows organization of the electron transfer chain of PSII in a pseudo-C2 symmetry by two subunits D1 and D2. The water-oxidation reaction proceeds via the Si-state cycle 8 (with i=0-4), where dioxygen is produced in the transition of S3→(S4)→S0 (Fig. 1b). The high-resolution structures of PSII analyzed so far were for the dark-stable S1 state 4,5 , although a few studies on the low-resolution intermediate S-state structures have been reported by TR-SFX [9][10][11] . During the revision of our manuscript, Young et al. reported a 2F-illuminated state structure at 2.25 Å resolution where no apparent changes around O5 were observed 12 , although estimations of the resolution could yield somewhat different values so that small movement of some water molecules may escape the detection. In order to achieve resolution high enough to uncover small structural changes induced by flash illuminations yet allowing Si-state transition to proceed efficiently, we determined the optimal crystal size of PSII with a maximum length of 100 µm, which diffracted up to a resolution of 2.1 Å by a SACLA-XFEL ...

The photosynthetic apparatus of green sulfur bacteria (GSB) contains a peripheral antenna chlorosome, light-harvesting Fenna-Matthews-Olson proteins (FMO), and a reaction center (GsbRC). We used cryo–electron microscopy to determine a 2.7-angstrom structure of the FMO-GsbRC supercomplex from Chlorobaculum tepidum. The GsbRC binds considerably fewer (bacterio)chlorophylls [(B)Chls] than other known type I RCs do, and the organization of (B)Chls is similar to that in photosystem II. Two BChl layers in GsbRC are not connected by Chls, as seen in other RCs, but associate with two carotenoid derivatives. Relatively long distances of 22 to 33 angstroms were observed between BChls of FMO and GsbRC, consistent with the inefficient energy transfer between these entities. The structure contains common features of both type I and type II RCs and provides insight into the evolution of photosynthetic RCs.

Researchers are still discussing the classification of Nycticebus. We established a molecular phylogeny covering all recognized taxa in Nycticebus to provide information for further evaluation. We sequenced partial D-loop (ca. 390 bp) and cytochrome b genes (425 bp) from 22 specimens. We separated most of the major groups except for some mixing of Nycticebus. coucang coucang and N. bengalensis. Nycticebus pygmaeus diverged earlier from the ancestral stock than the other taxa. Nycticebus coucang menagensis was well discriminated from N. c. coucang. It may be possible to explain the mixing of Nycticebus coucang coucang and N. bengalensis by the hybridization between the 2 groups in the field. Although our data did not provide direct 1188 Chen et al. evidence for or against the new proposal that Nycticebus coucang javanicus be raised to the rank of a distinct species (N. javanicus), we have good evidence for regarding N. c. menagensis as a species.

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Bengal slow lorises (Nycticebus bengalensis) and pygmy slow lorises (Nycticebus pygmaeus) are nocturnal which creates difficulties to study them in the field. There is a scarcity of data on them and their population genetics are poorly understood. We sequenced and analyzed a partial fragment in the first hypervariable region of the mitochondrial control region or D-loop HVRI of 21 Nycticebus bengalensis and 119 N. pygmaeus from the boundary between China and Vietnam where they are sympatric. Though the sample size for Nycticebus pygmaeus is much larger, the polymorphism level is much lower than that of N. bengalensis, possibly Int J Primatol (2007) 28:791-799

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The UV-Vis absorption spectrum of kaempferol and DPPH was investigated, the optimum determination wavelength and reaction time for determining the DPPH radical scavenging activity of kaempferol was 517 nm and 30 min, respectively. Kaempferol exhibited strong DPPH radical scavenging activity with a IC50 value of 0.004349 mg·mL-1, which is smaller than that of rutin, indicating that kaempferol has a stronger antioxidant activity than rutin.

The thermophilic purple sulfur bacterium Thermochromatium tepidum possesses four main water-soluble redox proteins involved in the electron transfer behavior. Crystal structures have been reported for three of them: a high potential iron-sulfur protein, cytochrome c', and one of two low-potential cytochrome c (which is a flavocytochrome c) have been determined. In this study, we purified another low-potential cytochrome c (LPC), determined its N-terminal amino acid sequence and the whole gene sequence, characterized it with absorption and electron paramagnetic spectroscopy, and solved its high-resolution crystal structure. This novel cytochrome was found to contain five c-type hemes. The overall fold of LPC consists of two distinct domains, one is the five heme-containing domain and the other one is an Ig-like domain. This provides a representative example for the structures of multiheme cytochromes containing an odd number of hemes, although the structures of multiheme cytochromes with an even number of hemes are frequently seen in the PDB database. Comparison of the sequence and structure of LPC with other proteins in the databases revealed several characteristic features which may be important for its functioning. Based on the results obtained, we discuss the possible intracellular function of this LPC in Tch. tepidum.