Search

Purpose Few studies have compared the implant quality of linked and loose seeds for prostate brachytherapy. This study aimed to evaluate and compare plan reproducibility of intraoperatively built custom linked seeds and loose seeds for prostate brachytherapy. Material and methods Between December 2010 and March 2014, 76 localized prostate cancer patients received Iodine-125 brachytherapy with external beam radiotherapy. Linked and loose seeds were implanted in 39 and 37 patients, respectively. The primary endpoint was the mean (± standard deviation) of the absolute change in the minimum dose received by 90% of the prostate volume between intraoperative and post-operative planning (ΔD90) to confirm plan reproducibility. Comparisons between the groups were evaluated using 2-sample t tests. Results The ΔD90 values were 6.95 ± 11.6% and -0.41 ± 8.5% for the loose and linked seed groups, respectively (p < 0.01). The linked seed group showed decreased post-operative D90 (118.8% vs. 127.2%), V150 (51.7% vs. 66.7%), and RV100 (0.44 ml vs. 0.61 ml) compared to the loose seed group (p < 0.01), whereas lung migration tended to be reduced (0% vs. 8%). Conclusions The plan reproducibility of the linked seed group was better than that of the loose seed group. Moreover, the linked seed group showed less migration and lower rectal dose.

Purpose To analyze genitourinary toxicity by followup of the International Prostate Symptom Score (IPSS) and Overactive Bladder Symptom Score (OABSS) after prostate brachytherapy. Methods and Materials Six hundred eighty patients were treated with iodine-125 brachytherapy for localized prostate cancer. IPSS, OABSS, and two categories of IPSS questions (storage symptom score [IPSS-S] and voiding symptom score [IPSS-V]) were evaluated. Results The median followup was 54 months (range, 24–108). All scales showed rapid increases followed by gradual decreases. The median times to IPSS peak and resolution were 1 and 6 months, respectively. The resolution rates of IPSS, IPSS-S, IPSS-V, and OABSS at the last followup were 84.2%, 86.3%, 89.5%, and 83.0%, respectively. The difference between IPSS baseline and peak was greater for larger preimplant prostate volumes (≥25 mL, p = 0.004). The time to resolution was longer for higher biologic effective dose (BED) (≥210 Gy, p = 0.019 [IPSS]), in those with larger prostate volumes (≥25 mL, p = 0.025 [OABSS]), in younger patients (younger than 70 years, p = 0.043 [IPSS-S]), and in those with androgen deprivation therapy (ADT) use ( p = 0.049 [IPSS-V]). Urge incontinence, included in the OABSS, was observed more commonly in older patients (75 years and older, p = 0.018), with ADT use ( p p = 0.006). Conclusions The IPSS and OABSS showed similar patterns of change. Urinary symptoms improved more rapidly in those with high baseline IPSS levels. The OABSS was useful for following urinary symptoms after prostate brachytherapy. Age, ADT use, preimplant prostate volume, and BED were significantly associated with urinary outcomes.

Purpose To compare late genitourinary (GU) and gastrointestinal (GI) toxicity following different prostate cancer treatment modalities. Materials and methods This study included 1084 consecutive prostate cancer patients treated with conventional radiotherapy, intensity-modulated radiotherapy (IMRT), permanent iodine-125 implantation (PI) alone, and PI combined with external beam radiotherapy (PI+EBRT). The effects of treatment- and patient-related factors on late grade⩾2 (G2+) GU/GI toxicity risk were assessed. Results The median follow-up was 43months (range, 12–97months). Compared to the PI+EBRT, there was significantly less G2+ GU toxicity in the conventional radiotherapy (hazard ratio [HR]=0.39; 95% CI, 0.20–0.77) and the IMRT (HR=0.45, 95% CI, 0.27–0.73). Compared to the PI+EBRT, there was significantly more G2+ GI toxicity in the IMRT (HR=2.38; 95% CI, 1.16–4.87). In PI-related groups, prostate equivalent dose in 2Gy fractions was a significant predictor of G2+ GU toxicity ( p =0.001), and the rectal volume receiving more than 100% of the prescribed dose was a significant predictor of G2+ GI toxicity ( p =0.001). Conclusion The differences in the late G2+ GU/GI risk cannot be explained by the differences in treatment modalities themselves, but by the total radiation dose to the GU/GI tract, which had a causal role in the development of late G2+ GU/GI toxicity across all treatment modality groups.

The archaeal toxin, aRelE, in the hyperthermophilic archaeon Pyrococcus horikoshii OT3 inhibits protein synthesis, whereas its cognate antitoxin, aRelB, neutralizes aRelE activity by forming a non-toxic complex, aRelB–aRelE. The structural mechanism whereby aRelB neutralizes aRelE activity was examined by biochemical and biophysical analyses. Overexpression of aRelB with an aRelE mutant (ΔC6), in which the C-terminal residues critical for aRelE activity were deleted, in Escherichia coli allowed a stable complex, aRelB–ΔC6, to be purified. Isothermal titration of aRelE or ΔC6 with aRelB indicated that the association constant ( K a) of wild-type aRelB–aRelE is similar to that of aRelB–ΔC6, demonstrating that aRelB makes little contact with the C-terminal active site of aRelE. Overexpression of deletion mutants of aRelB with aRelE indicated that either the N-terminal (pos. 1–27) or C-terminal (pos. 50–67) fragment of aRelB is sufficient to counteract the toxicity of aRelE in E. coli cells and the second α-helix (α2) in aRelB plays a critical role in forming a stable complex with aRelE. The present results demonstrate that aRelB, as expected from its X-ray structure, precludes aRelE from entering the ribosome, wrapping around the molecular surface of aRelE.

Time-resolved fluorescence spectroscopy of tryptophan residue has been extensively applied to the studies on structure-function relationships of protein. Regardless of this, the fluorescence decay mechanism and kinetics of tryptophan residue in many proteins still remains unclear. Previous studies have demonstrated that conformational heterogeneity and relaxation dynamics are both involved in the peculiar multiexponential decay kinetics in subnanosecond resolution. In the present study, we characterized the fluorescence decay property of six indole compounds in glycerol by resolving the contribution of conformational heterogeneity and relaxation dynamics. We applied the time-resolved area-normalized fluorescence emission spectrum (TRANES) method for the fluorescence decay analysis. The results of TRANES, time-dependent shift of fluorescence spectral center of gravity, and fluorescence decay simulation demonstrated that the dielectric relaxation process independent of intrinsic rotamer/conformer and the individual fluorescence lifetime gives the peculiarity to the fluorescence decay of indole compounds. These results confirmed that TRANES and time-dependent spectral shift analysis are potent methods to resolve the origin of multiexponential decay kinetics of tryptophyl fluorescence in protein.

The steady-state and time-resolved fluorescence spectroscopy is one of the most powerful method to detect and analyze subtle conformation change and interaction between peptide elements in protein. Phytocystatin Scb isolated from sunflower seeds includes a single Trp residue at position 85. In an attempt to investigate the interaction of the N-terminal region of Scb with the first and second hairpin loops by fluorescence spectroscopy of Trp residue, two Scb mutants in which single Trp locates at position 52 and 58, respectively, and their N-terminal removed mutants were generated. The N-terminal truncation changed the fluorescence decay kinetics of Trp52 from the triple exponential to double. Furthermore, the time-resolved fluorescence anisotropy residue indicated that the segmental motion of Trp52 was significantly enhanced by its N-terminal truncation. In contrast, Trp58 and Trp85 had little influence. The N-terminal successive truncations of Scb and its mutants resulted in the weaken inhibitors to papain. These results suggested that the N-terminal region of Scb interacts with the peptide segment preceding the first hairpin loop, thereby stabilizing the conformation of the hairpin loop structure.

The conformation and dynamics of a protein are essential in characterizing the protein folding/unfolding intermediate state. They are closely involved in the packing and site-specific interactions of peptide elements to build and stabilize the tertiary structure of the protein. In this study, it was confirmed that trypsin inhibitor obtained from seeds of bitter gourd (BGTI) adopted a peculiar but plausible conformation and dynamics in the unfolding intermediate state. The fluorescence spectrum of one of two tryptophan residues of BGTI, Trp9, shifted to the blue side in the presence of 2-3 M guanidine hydrochloride, although the other, Trp54, did not show this spectral shift. At the same time, the motional freedom of Trp9 revealed by a time-resolved fluorescence study decreased, suggesting that the segmental motion of this residue was more restricted. These results indicate that BGTI takes such a conformation state that the hydrophobic core and loop domains arranging Trp9 and Trp54 respectively are heterogeneously packed in the unfolding intermediate state.

Intermolecular interactions between protein molecules diffusing in various environments underlie many biological processes as well as control protein crystallization, which is a crucial step in x-ray protein structure determinations. Protein interactions were investigated through protein rotational diffusion analysis. First, it was confirmed that tetragonal lysozyme crystals containing fluorescein-tagged lysozyme were successfully formed with the same morphology as that of native protein. Using this nondisruptive fluorescent tracer system, we characterized the effects of sodium chloride and ammonium sulfate concentrations on lysozyme-lysozyme interactions by steady-state and time-resolved fluorescence anisotropy measurements and the introduction of a novel interaction parameter, krot. The results suggested that the specific attractive interaction, which was reflected in the retardation of the protein rotational diffusion, was induced depending on the salt type and its concentration. The change in the attractive interactions also correlated with the crystallization/precipitation behavior of lysozyme. Moreover, we discuss the validity of our rotational diffusion analysis through comparison with the osmotic second virial coefficient, B22, previously reported for lysozyme and those estimated from krot.

Snowmelt runoff is a precious water resource in cold region. However it causes disasters by influence of a warm air and a rainfall in spring. Therefore accurate estimation of snow water equivalent and forecast of snowmelt discharge are vital for river management. We investigated how snow water equivalent changes with time in the Jozankei Dam basin and on the premises of the Ishikari Experiment Laboratory. It was done by continuously monitoring of snowcover weight and snowmelt discharge using snowcover weightmeters and lysimeters. We observed time-lag of snowmelt runoff and excessive snowmelt runoff. It was caused by structure of snowcover layer. We proposed a simple equation for estimation of snow density by multiple linear regression analysis. We estimated snow water equivalent in the Jozankei dam basin by using the equation. The result of estimation is almost satisfactory.

Field observation of turbidity, chlorophyll and current velocity was conducted in the Barato River, which is the oxbow lake in the Ishikari River. In this oxbow lake resuspension of the sediment is main factor of water pollutant. In this survey, relationship among sediment resuspension, wind speed and current velocity were investigated. And the rate of sediment resuspesion was estimated by using observation results.Clear relationship exists between wind speed and turbidity when the wind speed was larger than certain level in each survey point. We tried to estimate the rate of sediment resupension by the methods in our study. The estimated value was generally adjusted to observed results in 2005; however, considerable difference was found in 2006.

The SOFC cell-stacks called flat-tube segmented-in-series cell-stacks have been developed by a joint work of Tokyo Gas and Kyocera. The cell-stacks were composed of multiple single- cells (8YSZ electrolyte, Ni/8YSZ anode, and LaSrCoFe cathode) and current-collectors, placed on both sides of the flat- tubular substrate. The fuel to electric power-generation efficiency of the latest cell-stack converted as the methane fuel was estimated more than 53% HHV-DC at 1023K at the current density, 0.2A/cm2. The complex impedance spectra of the cell- stacks were measured under the operating conditions, for investigations of the relation between the performance and components design. By the measurements under several conditions, we identified the relationship between the impedance spectra and the fuel usage dependence and degradation of the segmented-in-series type cell-stack performance. Based on these results, we continue further studies of the component design (especially anode), for higher efficiency and long-term durability of the cell-stacks.

Approximately half of the precipitation in Hokkaido falls as snow, and snowmelt floods in spring are an important factor in the river environment and in flood control and other river management. In Hokkaido, water storage at dams is managed using snowmelt water, and accurate estimation of snow water equivalent is important for flood management. We investigated how snow water equivalent changes with time in the Jozankei Dam basin, a basin that typifies those in snowy regions. Investigation was done by continuously monitoring snowcover weight and melted weight using a snowcover weightometer, which allows direct measurement of snowcover weight, and other instruments. Temporal changes in snow water equivalent and snow density were identified. The snow density was around 0.2g/cm3 at the initial stage of snow-accumulation, but at the late stage, the snow became granular in all layers and its density stabilized at around 0.4-0.5 g/cm3.

This study evaluates the living environment for Oncorhynchus masou parr by their feeding activities. Knowledge on physiology and ecology of Oncorhynchus masou parr is used for calculation of acquired energy after feeding, and consumed energy for maintaining their position and resting against current. The balance between the two kinds of energy is also calculated. The evaluation result can be applied for relative evaluation of the living environment of each section that show different environmental data, and for quantitative estimation of fish habitation in other rivers. To support this, the study verifies the validity of the evaluation by analyzing pools and riffles in the studied sections of the river, and the relationship between the energy balance and fish population in the river channel. And we clarified the optimal environmental conditions for Oncorhynchus masou.

Field test results of residential SOFC co-generation system at Tokyo Gas T. Ishikawa, S. Yamashita Product Development Dept., Tokyo Gas Lo., LTD., Tokyo, JAPAN We are developing residential SOFC co-generation system in order to commercialize it. One of our activities is to operate SOFC systems in field tests to make sure effects of installation and find technical issues. The detail is as below. A serious accident at nuclear power plants in Fukushima which was triggered by a huge earthquake on 11th March 2011 forces drastic re-formulation of Japan’s energy policy. In a series of discussions on the new energy policy, promoted use of distributed power generation is demanded more than ever as one of the main power systems, with extremely strong expectations for the fuel cell co-generation system. Tokyo Gas has been conducting research and development of solid oxide fuel cell (SOFC) which is expected to achieve the highest efficiency among all distributed power systems. In parallel with it, to verify the applicability of the SOFC for residential use and to identify technical challenges of SOFC, we have started field tests of SOFC systems of multiple manufacturers in the “Solid Oxide Fuel Cell (SOFC) demonstrative research project” with financial support from NEDO (Figure 1) [1][2]. We have installed a total of 23 units of four manufacturers in the field test of over four years (from 2007 to 2010), of which 14 units are still under operation as of April 30, 2012 (table 1). And the operating time has reached 18,600 hours at the longest, with 268,500 hours in total of 23 units. We have acquired the knowledge as shown below during the field tests conducted at general residences which we selected to reflect a variety of electrical and hot water demands (Figure 2). (1) Residential SOFC co-generation system contributes to the saving the preliminary energy consumptions and CO2 emissions by households in Japan (Figure 3, Figure 4). (2) SOFC system shows high electricity efficiency exceeding 45%LHV, and the transition of efficiency indicates that the durability of SOFC system including cell stack is estimated to be almost practical use (Figure 5). (3) SOFC system shows high reliability because of its simple system configuration. We will report the detailed results of SOFC field tests based on the latest data. Acknowledgments Finally, we would like to express our appreciation to NEDO, as this report is prepared mainly based on the results of the project supported by NEDO. Figure 2 Residential Demand (/Month) References [1] Solid Oxide Fuel Cell demonstrative research project final report (2011) http://www.nef.or.jp/sofc/share/pdf/h22y.pdf [2] Y. KAWABATA, T.SEYAMA, T.SOBUE, International Gas Union Research Conference ( 2011 ) Development and Field Test of Residential SOFC Co-generation System with All-ceramics-segmented Cells

The ribonuclease P (RNase P) in the hyperthermophilic archaeon Pyrococcus horikoshii comprises RNA (PhopRNA) and five proteins. We analyzed the RNA binding mode of the protein, using a pair of complementary fluorescence-labeled oligoribonucleotides. Fluorescence resonance energy transfer (FRET)-based assays suggested that the RNase P proteins assist PhopRNA in attaining a functionally active conformation via a distinct mode of binding.

The spectroscopic parameters of 4,7-bis(2-thienyl)-1,2,5-oxadiazolo[3,4- c ]pyridine (DTOP) were determined which was designed as a red-emitting dye for the electroluminescent (EL) device. The main optical transition of DTOP is attributable to 1 L a , and therefore, the fluorescence maximum shifted to the red side to reach to 630 nm according to the solvents. Although the non-radiative transition rate was enhanced, when the fluorescence maximum shifts to the longer than 610 nm, DTOP maintains the higher radiation transition probability.

A de novo designed pore-forming small globular protein (SGP) with antitumor activity consists of four helices: 3 basic amphipathic helices composed of Leu and Lys surrounding a central hydrophobic helix composed of oligoalanine. These helices are connected by a β-turn-forming sequence and two β-turn-unfavorable ones (S. Lee, T. Kiyota, T. Kunitake, E. Matsumoto, S. Yamashita, K. Anzai, and G. Sugihara Biochemistry 1997, Vol. 36, pp. 3782–3791). In the present work, we designed and synthesized three new SGP analogs in order to study the stoichiometric packing geometry and stability of SGP. The replacement of alanines in the central helix of SGP with leucines (SGP-L), which make the helix much larger in size and more hydrophobic, resulted in an equilibrium of monomeric–trimeric structure. The replacement of some Lys residues by Glu residues in the hydrophilic regions of the amphipathic helices (SGP-E) led to a decrease in helical content and the formation of an equilibrium of monomeric–trimeric structure. The alteration of β-turn regions with Gly residues, which makes these regions flexible (SGP-G), established an equilibrium of monomeric–dimeric states in buffer. The hydrophobic α-helix of SGP-L penetrated into the lipid bilayers in a manner that stabilized model membranes and biomembranes, whereas the central helices of SGP-G and -E destabilized them by forming channels. SGP and its analogs may be a useful model to study the role of the hydrophobic and hydrophilic regions in the formation of monomer–oligomer of proteins and to better understand the insertion of membrane targeting proteins into biomembranes. © 2001 John Wiley & Sons, Inc. Biopolymers 56: 96–108, 2001

The fluorescence spectral distributions of four tryptophan residues of hen egg-white lysozyme were analyzed using time-resolved and quenching-resolved fluorescence spectroscopy. Trp62 and Trp108 gave the fluorescence maxima at 352 nm and 342 nm, respectively. The fluorescence of Trp28 and Trp111 occurred only at 300-360 nm and they were observed as an unresolved emission band with a maximum and shoulder at 320 nm and 330 nm. The fluorescence quenching and decay parameters of each tryptophan residue reconfirmed that Trp62 was fully exposed to the solvent but Trp108 was sealed in the cage of the peptide chains and furthermore showed that Trp28 and Trp111 are under the influence of the larger fluctuational motion at the hydrophobic matrix box. The fluorescence responses of each tryptophan residue to the lysozyme-ligand interaction suggested that the internal fluctuation was reduced by the binding of ligand to give a distorted conformation to the hydrophobic matrix box region.

The mode of interaction of two pyrenylalanine residues which are introduced at positions 1 and 3′ into gramicidine S, [Pya 1,3′ ]GS, was investigated through absorption, CD, and steady-state and time-resolved fluorescence spectroscopy in organic and buffer solutions. CD spectra showed that the pyrene moieties could adopt an orientation to form a sandwich-type close pair on the two strands in a typical antiparallel β-structure in trifluoroethanol. Other spectroscopic studies demonstrated that the pyrenes of [Pya 1,3′ ]GS in trifluoroethanol formed specific intramolecular association on the pyrene pair, a ‘sandwich-type dimer’, in the ground state.

Tachyplesin I, an antibacterial and antiviral heptadecapeptide from the hemocyte debris of Tachypleus tridentatus, contains four aromatic amino acids (Trp2, Phe4, Tyr8, and Tyr13) which have been shown to be crucial for activity. In order to investigate conformations and orientations of the aromatic amino acid residues of tachyplesin I in lipid bilayers, its analogs, [Phe8]- and/or [Phe13]-tachyplesin(s) I in which Tyr8 and Tyr13 are replaced by Phe, were synthesized. Circular dichroism spectral studies showed that three peptides are considerably different in conformation in aqueous solution at pH 8.0 whereas they take similar conformations in the presence of neutral egg yolk phosphatidylcholine (EYPC) liposomes. Energy transfer kinetics showed that the distances of Trp2-Tyr8 and Trp2-Tyr13 are 16 A (max of 18.3 A, min of 15.1 A) and 18 A (max of 20.2 A, min of 16.6 A) in buffer but are 12 A (max of 15.2 A, min of 8.6 A) and 18 A (max of 22.9 A, min of 12.9 A), respectively, in the presence of acidic EYPC/EYPG (3:1) liposomes. Decay kinetics for Trp2 fluorescence indicated that Trp2 takes at least three conformations in buffer and in acidic liposomes where fractions of three Trp2 conformers vary by changing the medium from buffer to acidic liposomes. Although tachyplesin I is not in amphiphilic structure in buffer in spite of its rigid antiparallel beta-conformation, its interaction with lipid bilayers appears to induce amphiphilic structure via minor alteration of peptide backbone and side chain orientations, resulting in shortening the distance of Trp2-Tyr8.

In Escherichia coli the oriC-independent primosome plays an essential role in replication restart after dissociation of the replication DNA–protein complex by DNA damage. Primosome is thought to form via two pathways: one PriA dependent and the other PriA independent. PriC is a key protein in the replication restart of the PriA-independent pathway. In this study, we determined that PriC was divided into two domains. Then, we obtained information that: (i) the C-terminal domain preferentially binds to single-stranded DNA (ssDNA); (ii) the binding of PriC to ssDNA depends on salt concentration; and (iii) the binding site size of PriC is approximately 7–9 nucleotides. The protease digestion of PriC suggested that a possible DNA-binding site is the N-terminus of the C-terminal domain where basic amino acid residues are concentrated. Interestingly, α-helical induction of the C-terminal domain of PriC occurred after the addition of DNAs. Also, we examined the role of heptad repeat of leucine or valine residues in the C-terminal domain and PriC oligomerization. This study describes the structure and function analysis of PriC which forms the primosome complex in replication restart.

The cationic derivatives, the hydrochloride salts of ethylenediamine monodeoxycholate (EdaDC) and cholate (EdaC) derived from the respective bile salts, sodium deoxycholate (NaDC) and cholate (NaC), were synthesized and their physicochemical and biological properties together with the possibility of application as crystallizing agents for membrane proteins were studied. The critical micelle concentrations (CMCs) in water were determined to be 4.0 mM for EdaDC and 9.0 mM for EdaC, and the surface tensions were lowered down to 43 and 41 mN m−1, respectively, in the range above CMCs. When compared with the corresponding natural bile salts, the two cationic bile acid derivatives have the highest surfactant ability, followed by the corresponding taurine-conjugated salts of TDC (taurodeoxycholic acid) and TC (taurocholic acid) and the free bile acid salts of DC and C have the lowest activity. Studies of dye release from pre-encapsulated vesicles, hemolysis and antimicrobial activity indicated that the concentrations leading to a large increase in the membrane permeability and biomembrane perturbation ability are signiicantly lower than the CMCs of these surfactants. Buffer solution containing EdaC or EdaDC could solubilize a membrane protein, light-harvesting chlorophyll ab protein complex (LHCII), at concentrations higher than CMCs, but the power was estimated to be one third of that of surfactants such as Triton X-100, n-octyl-β-d-glucopyranoside and n-nonyl-β-d-glucopyranoside. Fluroescence depolarization studies showed that some specific molecular interactions between LHCIIs are induced by the cationic bile salts, suggesting that the latter would be valid for the crystallization of LHCII. In fact, LHCII crystals were observed in crystallization media containing 1% EdaC or 0.5% EdaDC.

The ligand-induced conformational change of an active lysozyme derivative, Kyn62-lysozyme, in which Trp62 of hen egg-white lysozyme (EC 3.2.1.17) was selectively modified to kynurenine, was investigated by steady-state and time-resolved fluorescence spectroscopy. Kyn62 formed an intramolecular energy transfer donor-acceptor pair with a tryptophan residue as a donor. The energy transfer was related to the conformation of the active site. The spectral overlap integral (J) of the kynurenine-tryptophan pair is large as it was determined to be 4.92 x 10(-15) M-1 cm3. Time-resolved fluorescence properties of Kyn62-lysozyme and its complex with a trimer of N-acetyl-D-glucosamine [(GlcNAc)3] show that the energy donor is Trp28 or Trp111 in the hydrophobic matrix box of the free Kyn62-lysozyme. In the complex, it appears that the kynurenine residue drastically changed its orientation or approached closer to Trp108 to accept more efficiently the excitation energy from Trp108 on the binding of Kyn62-lysozyme with (GlcNAc)3.

The hydraulic characteristics of an ice-covered river is much different from those at other seasons, because the frictional resistance between water surface and icesheet increases due to the ice cover on the water surface.Accuracy of estimation of the average velocity is dependent on the velocity profile. Usually, the k-e model is used to estimate the velocity profile under ice cover. This paper verifies a new estimation of a velocity profile under ice, the Two-Power LAW, using data measured in Hokkaido.

The structural feature of unfolding intermediate of pokeweed anti-viral protein (PAP) was characterized using time-resolved fluorescence spectroscopic methods to elucidate protein folding/unfolding process. CD and fluorescence spectra consistently demonstrated that the unfolding of PAP completed at 4 M of guanidine hydrochloride (GuHCl). The fluorescence resonance energy transfer (FRET) and time-resolve fluorescence depolarization analysis of Trp208 and Trp237 located in the C-terminal domain of PAP suggested that peculiar unfolding intermediate populated before reaching to the unfolding state. The FRET distance of Trp237 to Tyr182 was extended to more than 28 A with keeping the compact conformation in the unfolding intermediate state populated in the presence of 2 M GuHCl. On the other hand, Trp208 maintained the energy transfer pair with Tyr72 near the active site, although the rotational freedom was increased a little. There results suggest that the most distinguished structural feature of the unfolding intermediate of PAP is the separation of C-terminal domain from N-terminal domain. FRET and fluorescence depolarization studies also showed that C-terminal domain would be more separated to liberate the segmental motions of Trp208 and Trp237 distinctly at the unfolding state.

Protein intermolecular depletion interactions induced by polyethylene glycol (PEG) depend largely on its concentration and molecular weight. Herein, we investigated the effects of various concentrations and molecular weights of PEG on lysozyme interactions through the analysis of protein rotational diffusion, which is susceptible to intermolecular interactions at short range. To this end, we measured fluorescence anisotropy of fluorescein-tagged lysozyme added as a tracer in concentrated native lysozyme solutions and introduced a protein concentration-dependent interaction parameter, k(rot). The results show the nonmonotonic changes in k(rot) as the concentrations of PEG10000 and 6000 are increased. The depletion attractions are characterized by the decrease in k(rot), indicating an increase of a degree at which protein rotational diffusion slows down. The influences of temperature on the lysozyme rotational diffusion and k(rot) were also measured, and the validity of this approach was checked through comparison with the colloidal theory.

Purpose To compare contrast-enhanced T1-weighted (CE-T1WI) magnetic resonance imaging (MRI) with computed tomography (CT) for postimplant dosimetry and seed recognition in prostate brachytherapy. Methods and Materials A total of 245 patients who received 125 I prostate brachytherapy with or without external beam radiotherapy were enrolled. For postimplant analysis, CT and MRI scans were obtained at 1 month after seed implantation. For MRI-based dosimetry, T2-weighted images were fused with the CE-T1WI; the prostate was delineated on the T2-weighted images, and the seed detection was performed manually on the CE-T1WI. In CT-based dosimetry, the seed detection was essentially performed automatically. The dosimetric results obtained by MRI-based and CT-based dosimetry were compared. Results The mean prostate D 90 (the minimum dose received by 90% of the prostate volume) estimated by MRI-based and CT-based dosimetry were 113% and 115%, respectively, with no significant difference. The mean prostate V 100 (the percent volume of the postimplant prostate receiving 100% of the prescribed dose) estimated by MRI-based and CT-based dosimetry were 95.2% and 95.8%, respectively, again with no significant difference. The mean prostate V 150 (the percent volume of the postimplant prostate receiving 150% of the prescribed dose) estimated by MRI-based and CT-based dosimetry were 52.8% and 57.0%, respectively ( p V 150 were at least 10% lower than the CT-based results, the seed detection by CT-based dosimetry was overestimated in highly seed-clustered areas or in the areas close to calcifications because of reconstruction artifacts in CT images. Conclusions MRI-based dosimetry using CE-T1WI appears to be acceptable. Our results suggest that MRI-based dosimetry is a practical method for estimation of the higher dose distribution, especially if seeds are clustered together or when they are close to calcifications.

Pyruvate dehydrogenase multienzyme complex was purified from Bacillus caldolyticus. The complex was composed of four polypeptides with molecular masses of 39.8, 41.7, 53.7, and 57.5kDa estimated by SDS-PAGE and they were presumed to be pyruvate decarboxylase (E1, dimeric), lipoate acetyltransferase (E2), and lipoamide dehydrogenase (E3) on the analogy of those from Bacillus stearothermophilus. E1 and E3 were stable at pH 5.7-10.2 and 4.5-11.3, respectively. Halves of E1 and E3 activity were abolished by incubation for 30min at 65°C and 85°C, respectively. Loss of overall activity was principally due to inactivation of E1. Structural changes in the complex incubated at high temperature were studied by fluorescence spectroscopy. The results suggested that the thermal denaturation of the complex proceeded through at least two different steps: inactivations of E1 and E3, and the former process is accompanied by a reduction of the complex size.

With reference to the local conformation of a protein, it is interesting to differentiate the individual fluorescence properties of included tryptophan residues without modification. The fluorescence spectrum of bitter gourd trypsin inhibitor (BGTI) was separated into two emission bands by the quenching-resolved fluorescence method. One emission band was given as a fraction with the Stern-Volmer quenching constant, 44.9 x 10(-3) M(-1), against the fluorescence quenching by KI, and it showed an emission maximum intensity at 341 nm. The fluorescence quenching constant of the other band was 1.58 x 10(-3) M(-1), and the maximum wavelength was found at 337 nm. These separated emissions were due to the fluorescence of Trp54 and Trp9 of BGTI. The quenching resolved-fluorescence spectrum was effectively applied to the precise description of the polar circumstances surrounding the Trp residues in the unfolding intermediate state of BGTI. The results suggested that the molten globule-like state of BGTI adopted such a peculiar conformation that the helix domain including Trp9 was packed more densely while the other loop domain partially unfolded.

Field measurements of velocity profiles for ice-covered streams were performed in Hokkaido. Surface conditions of the cover were classified into smooth ice cover, rippled ice cover and cover with frazil slush. Adaptability of logarithmic velocity distribution was first investigated. The friction factor fi and Manning's roughness coefficient ni were caluculated based on an assumption of the logarithmic distribution; average values ni for smooth and rippled cover were 0.012 and 0.021 respectively.

Human serum albumin (HSA) plays important roles in transport of fatty acids and binding a variety of drugs and organic compounds in the circulatory system. This protein experiences several conformational transitions by the change of pH, and the resulting conformations were essential for completing the physiological roles in vivo. Steady-state and time-resolved fluorescence spectroscopy was applied to single tryptophan residue solely arranged in HSA to study subtle conformational change around single tryptophan residue in HSA at various pH. The results showed the characteristic feature of local conformation around tryptophan residue in domain II responding to the change in entire structure. The study of time-resolved area-normalized fluorescence emission spectra (TRANES) also showed the peculiar dielectric property of water molecule trapped nearby tryptophan residue depending on pH. These results suggested that microenvironment around tryptophan residue was tightly packed at acidic and basic pH although entire conformation was loosened.

The unfolding of alpha-momorcharin was systematically investigated using steady-state and time-resolved tryptophan fluorescence, circular dichroism and 8-anilino-1-naphthalenesulfonic acid (ANS) binding. These spectroscopic studies demonstrated that alpha-momorcharin unfolded through a compact folded intermediate state. The content of alpha-helix was increased, Trp192 approached closer to the side of active site and its rotational motion was restricted by being equilibrated with 2-3 M of guanidine hydrochloride. Furthermore, the binding of ANS with alpha-momorcharin was more suppressed to show that the hydrophobic parts would not be accessed to the protein surface but rather be sealed off in this specific conformation state. These results suggest that the structure of alpha-momorcharin holds the more compact conformation as an incipient state for unfolding, which is the sharp contrast to beta-momorcharin that gives the characteristics of the generally known molten globule state.

Low-temperature pyrolysis of polycyclic aromatic quinones (dyes) has been attempted and the electrical conductivities of the pyrolyzed samples and their iodine-doped derivatives were checked. These samples will be of interest in possible applications as semiconductive materials in the electronics field.

The electronic structures of models for alkylene-aromatic polymers (AAPs) and their pyrolysis products, as member of the one-dimensional graphite family, have been studied on the basis of tight-binding crystal orbital calculations. The band gaps of AAPs decrease and the frontier-band widths increase with the progress of the pyrolysis. The change of the frontier electron distribution has also been examined.

The origin of multi-exponential fluorescence decay property of tryptophan (Trp) in protein has been in controversy, and dielectric relaxation is thought to be one of the most plausible candidates of that origin. In this study, we studied melittin-calmodulin interaction on the concept of dielectric relaxation by spectrally and time-resolved fluorescence spectroscopy. Trp residue in melittin demonstrated drastic change in its dielectric relaxation rate and scale by binding with calmodulin. Expected change of relaxation rate suggested that dielectric relaxation accounts for multi-exponential property of fluorescence decay. We also examined the time variation of radiative and non-radiative decay rates. That result demonstrated the distinct difference profiles of non-radiative decay rate of Trp in melittin and the complex.

The specific conformation of partially unfolded state of beta-momorcharin was characterized through the steady-state and time-resolved fluorescence spectroscopic studies on a single Trp-190 which located adjacently to the active site. The content of secondary structure was retained, the binding of ANS was remarkably enhanced, and the correlation time of entire protein rotation was prolonged at the partially unfolded state formed by being equilibrated with the mild concentration of guanidine hydrochloride. The time-resolved fluorescence depolarization and excitation energy transfer analysis suggest that Trp-190 approached 2 A closer to Tyr-70 and was hidden from the exposure to the protein surface, while the rotational correlation time and freedom of its segmental motion were shortened and enhanced, respectively. These results suggest that the transient folding/unfolding intermediate state of beta-momorcharin adopt the specific conformation at the vicinity of the active site, although it exhibits very similar properties with those of the generally known molten-globule state.

The thermal unfolding pathway for dihydrolipoamide dehydrogenase (LipDH) isolated from Bacillus stearothermophilus was investigated focusing on the transient intermediate state characterized through time-resolved fluorescence studies. The decrease in ellipticity in the far UV region in the CD spectrum, the fluorescence spectral change of Trp-91 and FAD, and the thermal enzymatic inactivation curve consistently demonstrated that LipDH unfolded irreversibly on heat treatment at higher than 65 degrees C. LipDH took a transient intermediate state during the thermal unfolding process which could refold back into the native state. In this state, the internal rotation of FAD was activated in the polypeptide cage and correspondingly LipDH showed a peculiar conformation. The transient intermediate state of LipDH characterized in time-resolved fluorescence depolarization studies showed very similar properties to the molten-globule state, which has been confirmed in many studies on protein folding.

The voltage loss in a single cell of a segmented-in-series flat-tube-type solid oxide fuel cell (SOFC) stack was evaluated. The stack exhibits a particular configuration, and the current flows along an in-plane direction in the anode and cathode; this differs from conventional planar-type SOFCs. Distributions of the current in the cell were studied in detail for many parameters such as the length of electrodes and conductivities of the cell components. Gas diffusion in a porous substrate and anode and the fuel utilization affect the concentration polarization; thus, they were also considered. By using the simulation results, appropriate configurations for the cell and stack were designed, and the simulation was validated by performance tests for a practical fuel cell stack. The stack exhibited a high performance of 0.76 V at 0.2 A/cm2 at Uf = 81.6% with dry H2 fuel, yielding a DC energy conversion efficiency of 53.6% HHV. Moreover, a bundle comprising 34 stacks was also fabricated, and excellent values of output voltage (342.5 V), output power (338.3 W), and electrical efficiency (49.5% HHV) at a current density of 0.2 A/cm2 were obtained.Copyright © 2006 by ASME