Hydrogen bonds of DsrD protein revealed by neutron crystallography

Hydrogen bonds of DNA-binding protein DsrD have been determined by neutron diffraction. In terms of proton donors and acceptors, DsrD protein shows striking differences from other proteins

Crystallization and preliminary X-ray studies of ferredoxin-NADP + oxidoreductase encoded by Bacillus subtilis yumC

金沢大学理工研究域物質化学系Ferredoxin-NADP+ oxidoreductase encoded by Bacillus subtilis yumC has been purified and successfully crystallized in complex with NADP + in two forms. Diffraction data from crystals of these two forms were collected at resolutions of 1.8 and 1.9 Å. The former belonged to space group P21212, with unit-cell parameters a = 63.90, b = 135.72, c = 39.19 Å, and the latter to space group C2, with unit-cell parameters a = 207.47, b = 64.85, c = 61.12 Å, Β = 105.82°. The initial structure was determined by the molecular-replacement method using a thioredoxin reductase-like protein as a search model. © 2010 International Union of Crystallography All rights reserved

Activation Process of [NiFe] Hydrogenase Elucidated by High-Resolution X-Ray Analyses: Conversion of the Ready to the Unready State

SummaryHydrogenases catalyze oxidoreduction of molecular hydrogen and have potential applications for utilizing dihydrogen as an energy source. [NiFe] hydrogenase has two different oxidized states, Ni-A (unready, exhibits a lag phase in reductive activation) and Ni-B (ready). We have succeeded in converting Ni-B to Ni-A with the use of Na2S and O2 and determining the high-resolution crystal structures of both states. Ni-B possesses a monatomic nonprotein bridging ligand at the Ni-Fe active site, whereas Ni-A has a diatomic species. The terminal atom of the bridging species of Ni-A occupies a similar position as C of the exogenous CO in the CO complex (inhibited state). The common features of the enzyme structures at the unready (Ni-A) and inhibited (CO complex) states are proposed. These findings provide useful information on the design of new systems of biomimetic dihydrogen production and fuel cell devices

An O-centered structure of the trinuclear copper center in the Cys500Ser/Glu506Gln mutant of CueO and structural changes in low to high X-ray dose conditions

Right on CueO: The O-centered structure of the trinuclear copper center in a multicopper oxidase (CueO) was shown to be an intermediate of the four-electron reduction of dioxygen (see picture). This structure was determined by in situ data collection of X-ray diffractions and copper K-edge spectra at low to high X-ray dose conditions. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

New insights into the catalytic active-site structure of multicopper oxidases

Structural models determined by X-ray crystallography play a central role in understanding the catalytic mechanism of enzymes. However, X-ray radiation generates hydrated electrons that can cause significant damage to the active sites of metalloenzymes. In the present study, crystal structures of the multicopper oxidases (MCOs) CueO from Escherichia coli and laccase from a metagenome were determined. Diffraction data were obtained from a single crystal under low to high X-ray dose conditions. At low levels of X-ray exposure, unambiguous electron density for an O atom was observed inside the trinuclear copper centre (TNC) in both MCOs. The gradual reduction of copper by hydrated electrons monitored by measurement of the Cu K-edge X-ray absorption spectra led to the disappearance of the electron density for the O atom. In addition, the size of the copper triangle was enlarged by a two-step shift in the location of the type III coppers owing to reduction. Further, binding of O2 to the TNC after its full reduction was observed in the case of the laccase. Based on these novel structural findings, the diverse resting structures of the MCOs and their four-electron O2-reduction process are discussed. © 2014 International Union of Crystallography

Upregulation of IGF-I in the goldfish retinal ganglion cells during the early stage of optic nerve regeneration

金沢大学医薬保健研究域　医学系Goldfish retinal ganglion cells (RGCs) can regrow their axons after optic nerve injury. However, the reason why goldfish RGCs can regenerate after nerve injury is largely unknown at the molecular level. To investigate regenerative properties of goldfish RGCs, we divided the RGC regeneration process into two components: (1) RGC survival, and (2) axonal elongation processes. To characterize the RGC survival signaling pathway after optic nerve injury, we investigated cell survival/death signals such as Bcl-2 family members in the goldfish retina. Amounts of phospho-Akt (p-Akt) and phospho-Bad (p-Bad) in the goldfish retina rapidly increased four- to five-fold at the protein level by 3-5 days after nerve injury. Subsequently, Bcl-2 levels increased 1.7-fold, accompanied by a slight reduction in caspase-3 activity 10-20 days after injury. Furthermore, level of insulin-like growth factor-I (IGF-I), which activates the phosphatidyl inositol-3-kinase (PI3K)/Akt system, increased 2-3 days earlier than that of p-Akt in the goldfish retina. The cellular localization of these molecular changes was limited to RGCs. IGF-I treatment significantly induced phosphorylation of Akt, and strikingly induced neurite outgrowth in the goldfish retina in vitro. On the contrary, addition of the PI3K inhibitor wortmannin, and IGF-I antibody inhibited Akt phosphorylation and neurite outgrowth in an explant culture. Thus, we demonstrated, for the first time, the signal cascade for early upregulation of IGF-I, leading to RGC survival and axonal regeneration in adult goldfish retinas through PI3K/Akt system after optic nerve injury. The present data strongly indicate that IGF-I is one of the most important molecules for controlling regeneration of RGCs after optic nerve injury. © 2007 Elsevier Ltd. All rights reserved

An Open-Label Feasibility Trial of Repetitive Transcranial Magnetic Stimulation for Treatment-Resistant Major Depressive Episodes

Repetitive transcranial magnetic stimulation (rTMS) has been reported to be a new treatment option for treatment-resistant depression. In Japan, there has been limited research into its feasibility, efficacy, and tolerability. We have launched a trial of rTMS for treating medication-resistant major depressive disorder and bipolar depression. We are investigating low-frequency rTMS to the right dorsolateral prefrontal cortex and traditional high-frequency rTMS to the left dorsolateral prefrontal cortex, in 20 patients. The primary outcome of the study is the treatment completion rate. This study will provide new data on the usefulness of rTMS for treatment-resistant depression in Japan