Despite the prevalence of obesity and its related diseases, the signaling pathways for appetite control and satiety are not clearly understood. Here we report C. elegans quiescence behavior, a cessation of food intake and movement that is possibly a result of satiety. C. elegans quiescence shares several characteristics of satiety in mammals. It is induced by high-quality food, it requires nutritional signals from the intestine, and it depends on prior feeding history: fasting enhances quiescence after refeeding. During refeeding after fasting, quiescence is evoked, causing gradual inhibition of food intake and movement, mimicking the behavioral sequence of satiety in mammals. Based on these similarities, we propose that quiescence results from satiety. This hypothesized satiety-induced quiescence is regulated by peptide signals such as insulin and TGF-beta. The EGL-4 cGMP-dependent protein kinase functions downstream of insulin and TGF-beta in sensory neurons including ASI to control quiescence in response to food intake.

The A2a adenosine receptor is a member of the G-protein coupled receptor family, and its activation stimulates cyclic AMP production. To determine the residues which are involved in ligand binding, several residues in transmembrane domains 5-7 were individually replaced with alanine and other amino acids. The binding properties of the resultant mutant receptors were determined in transfected COS-7 cells. To study the expression levels in COS-7 cells, mutant receptors were tagged at their amino terminus with a hemagglutinin epitope, which allowed their immunological detection in the plasma membrane by the monoclonal antibody 12CA5. The functional properties of mutant receptors were determined by measuring stimulation of adenylate cyclase. Specific binding of [3H]CGS 21680 (15 nM) and [3H]XAC (4 nM), an A2a agonist and antagonist, respectively, was absent in the following Ala mutants: F182A, H250A, N253A, I274A, H278A, and S281A, although they were well expressed in the plasma membrane. The hydroxy group of Ser-277 is required for high affinity binding of agonists, but not antagonists. An N181S mutant lost affinity for adenosine agonists substituted at N6 or C-2, but not at C-5'. The mutant receptors I274A, S277A, and H278A showed full stimulation of adenylate cyclase at high concentrations of CGS 21680. The functional agonist potencies at mutant receptors that lacked radioligand binding were > 30-fold less than those at the wild type receptor. His-250 appears to be a required component of a hydrophobic pocket, and H-bonding to this residue is not essential. On the other hand, replacement of His-278 with other aromatic residues was not tolerated in ligand binding. Thus, some of the residues targeted in this study may be involved in the direct interaction with ligands in the human A2a adenosine receptor. A molecular model based on the structure of rhodopsin, in which the 5'-NH in NECA is hydrogen bonded to Ser-277 and His-278, was developed in order to visualize the environment of the ligand binding site.

Human DJ-1 is a genetic cause of early-onset Parkinson's disease (PD), although its biochemical function is unknown. We report here that human DJ-1 and its homologs of the mouse and Caenorhabditis elegans are novel types of glyoxalase, converting glyoxal or methylglyoxal to glycolic or lactic acid, respectively, in the absence of glutathione. Purified DJ-1 proteins exhibit typical Michaelis-Menten kinetics, which were abolished completely in the mutants of essential catalytic residues, consisting of cysteine and glutamic acid. The presence of DJ-1 protected mouse embryonic fibroblast and dopaminergically derived SH-SY5Y cells from treatments of glyoxals. Likewise, C. elegans lacking cDJR-1.1, a DJ-1 homolog expressed primarily in the intestine, protected worms from glyoxal-induced death. Sub-lethal doses of glyoxals caused significant degeneration of the dopaminergic neurons in C. elegans lacking cDJR-1.2, another DJ-1 homolog expressed primarily in the head region, including neurons. Our findings that DJ-1 serves as scavengers for reactive carbonyl species may provide a new insight into the causation of PD.

The making and breaking of atomic bonds are essential processes in chemical reactions. Although the ultrafast dynamics of bond breaking have been studied intensively using time-resolved techniques, it is very difficult to study the structural dynamics of bond making, mainly because of its bimolecular nature. It is especially difficult to initiate and follow diffusion-limited bond formation in solution with ultrahigh time resolution. Here we use femtosecond time-resolved X-ray solution scattering to visualize the formation of a gold trimer complex, [Au(CN)2(-)]3 in real time without the limitation imposed by slow diffusion. This photoexcited gold trimer, which has weakly bound gold atoms in the ground state, undergoes a sequence of structural changes, and our experiments probe the dynamics of individual reaction steps, including covalent bond formation, the bent-to-linear transition, bond contraction and tetramer formation with a time resolution of ∼500 femtoseconds. We also determined the three-dimensional structures of reaction intermediates with sub-ångström spatial resolution. This work demonstrates that it is possible to track in detail and in real time the structural changes that occur during a chemical reaction in solution using X-ray free-electron lasers and advanced analysis of time-resolved solution scattering data.

The vibrational properties of the hydrated proton and deuteron in bulk phase water and deuterated water are investigated spectroscopically and computationally. Mid-infrared spectra of aqueous acid solutions are measured by attenuated total reflectance-Fourier transform IR spectroscopy and compared with pure water and salt/counterion spectra to extract high-quality hydrated proton spectra at a series of concentrations. Multistate empirical valence bond simulations of the excess proton in bulk phase water are also performed, allowing the autocorrelation function of the time derivative of the dipole moment, and hence the power spectrum of the hydrated proton, to be evaluated. The experimental and theoretical spectra are found to be in very good agreement. Normal mode analysis of the bulk phase simulation data allows definitive assignment of the spectrum. The associated motions are found to be represented by both Eigen and Zundel forms of the hydrated proton.

X-ray free electron lasers (XFELs) deliver short (<100 fs) and intense (∼10 12 photons) pulses of hard X-rays, making them excellent sources for time-resolved studies. Here we show that, despite the inherent instabilities of current (SASE based) XFELs, they can be used for measuring high-quality X-ray absorption data and we report femtosecond time-resolved Xray absorption near-edge spectroscopy (XANES) measurements of a spin-crossover system, iron(II) tris(2,2′-bipyridine) in water. The data indicate that the low-spin to high-spin transition can be modeled by single-exponential kinetics convoluted with the overall time resolution. The resulting time constant is ∼160 fs. ■ INTRODUCTIONChemical reactions involve the dynamic evolution of strongly coupled electrons and nuclei on ultrafast time scales from few to a few hundred femtoseconds. Disentangling this complex dynamics has motivated a wide range of experimental and theoretical studies to obtain a more complete picture of the elementary steps underlying these processes. 1 Despite the progresses in ultrafast optical spectroscopies that can resolve dynamics of the valence shell electrons with femtosecond time resolution and in theoretical calculations of many-body dynamics, our understanding of the early chemical transformations still remains incomplete due to the lack of direct structural measurements visualizing both electronic structure and nuclear movements in real time. Information on nuclear dynamics has been obtained with time-resolved scattering with a time resolution of 100 ps on a variety of system from solutions 2,3 to single crystals. 4−6 Recently, femtoseconds studies have been published. 7−9 When the surrounding of specific atoms is of interest, recent advances have demonstrated the utility of time-resolved X-ray absorption spectroscopy (XAS) as a reliable monitor of nuclear and electronic structure during chemical reactions on both the picosecond 10,11 and, more recently, the femtosecond time scale. 12−14 Yet, most of the synchrotron-based experiments are limited to insufficient time resolution (∼100 ps in normal operating conditions) or very low flux (∼10 photons/pulse at kilohertz rate, when operated in femtosecond laser slicing mode 15 ). The development of new X-ray facilities such as X-ray free electron lasers (XFELs) provide unprecedented capabilities in terms of both X-ray pulse durations (down to tens of femtoseconds) and single pulse intensities of more than 10 10 photons when a 1 eV bandwidth monochromator is used. In the present study we exploit XFEL radiation from the Linac Coherent Light Source (LCLS), the first hard XFEL, 16 and present an experimental study of the photoexcited spin transition dynamics in aqueous ferreous bipyridine ([Fe-(bpy) 3 ] 2+ ), which proves the feasibility of femtosecond XAS at the LCLS.Transition metal complexes exhibiting photoinduced spin crossover (SCO) transition provide a good example of the electronic dynamics coupled with nuclear motions. In a

Photoreceptor proteins play crucial roles in receiving light stimuli that give rise to the responses required for biological function. However, structural characterization of conformational transition of the photoreceptors has been elusive in their native aqueous environment, even for a prototype photoreceptor, photoactive yellow protein (PYP). We employ pump-probe X-ray solution scattering to probe the structural changes that occur during the photocycle of PYP in a wide time range from 3.16 μs to 300 ms. By the analysis of both kinetics and structures of the intermediates, the structural progression of the protein in the solution phase is vividly visualized. We identify four structurally distinct intermediates and their associated five time constants and reconstructed the molecular shapes of the four intermediates from time-independent, species-associated difference scattering curves. The reconstructed structures of the intermediates show the large conformational changes such as the protrusion of N-terminus, which is restricted in the crystalline phase due to the crystal contact and thus could not be clearly observed by X-ray crystallography. The protrusion of the N-terminus and the protein volume gradually increase with the progress of the photocycle and becomes maximal in the final intermediate, which is proposed to be the signaling state. The data not only reveal that a common kinetic mechanism is applicable to both the crystalline and the solution phases, but also provide direct evidence for how the sample environment influences structural dynamics and the reaction rates of the PYP photocycle.

Lactic acid bacteria are known to perform significant roles in the fermentation of kimchi, a fermented cabbage product. However, the microbial population dynamics inherent to kimchi fermentation remain to be clearly elucidated. In this study, we have characterized the microbial dynamics via the identification of a total of 970 bacterial isolates, representing 15 species of the genera Lactobacillus, Leuconostoc, and Weissella, all of which were primarily identified by PCR-based restriction enzyme analysis. These population dynamics appear to be influenced markedly by fermentation temperature. Distinct biphasic microbial growth was observed with preliminary 2-day incubation at 15 degrees C, conducted before main fermentation at -1 degrees C. Leuconostoc citreum, as well as Leuconostoc gasicomitatum, predominated during the first growth phase, whereas Weissella koreensis predominated during the second phase. By way of contrast, with preliminary 4-day incubation at 10 degrees C, only W. koreensis grew rapidly from the beginning of the process. Therefore, our findings suggest that a short incubation at 15 degrees C enhances the growth of the less psychrophilic Leuconostoc species, including Lc. citreum, thus delaying the growth of the predominant W. koreensis, which is a more adaptive species at -1 degrees C.