Heme Orientation of Cavity Mutant Hemoglobins (His F8 → Gly) in Either α or β Subunits: Circular Dichroism, 1H NMR,and Resonance Raman Studies

Native human adult hemoglobin (Hb A) has mostly normal orientation of heme, whereas recombinant Hb A (rHb A) expressed in E. coli contains both normal and reversed orientations of heme. Hb A with the normal heme exhibits positive circular dichroism (CD) bands at both the Soret and 260‐nm regions, while rHb A with the reversed heme shows a negative Soret and decreased 260‐nm CD bands. In order to examine involvement of the proximal histidine (His F8) of either α or β subunits in determining the heme orientation, we prepared two cavity mutant Hbs, rHb(αH87G) and rHb(βH92G), with substitution of glycine for His F8 in the presence of imidazole. CD spectra of both cavity mutant Hbs did not show a negative Soret band, but instead exhibited positive bands with strong intensity at the both Soret and 260‐nm regions, suggesting that the reversed heme scarcely exists in the cavity mutant Hbs. We confirmed by ¹H NMR and resonance Raman (RR) spectroscopies that the cavity mutant Hbs have mainly the normal heme orientation in both the mutated and native subunits. These results indicate that the heme Fe‐His F8 linkage in both α and β subunits influences the heme orientation, and that the heme orientation of one type of subunit is related to the heme orientation of the complementary subunits to be the same. The present study showed that CD and RR spectroscopies also provided powerful tools for the examination of the heme rotational disorder of Hb A, in addition to the usual ¹H NMR technique. Chirality 28:585–592, 2016. © 2016 Wiley Periodicals, Inc.     

Optical Activity Governed by Local Chiral Structures in Two‐Dimensional Curved Metallic Nanostructures

Chiral nanostructures show macroscopic optical activity. Local optical activity and its handedness are not uniform in the nanostructure, and are spatially distributed depending on the shape of the nanostructure. In this study we fabricated curved chain nanostructures made of gold by connecting linearly two or more arc structures in a two‐dimensional plane. Spatial features of local optical activity in the chain structures were evaluated with near‐field circular dichroism (CD) imaging, and analyzed with the aid of classical electromagnetic simulation. The electromagnetic simulation predicted that local optical activity appears at inflection points where arc structures are connected. The handedness of the local optical activity was dependent on the handedness of the local chirality at the inflection point. Chiral chain structures have odd inflection points and the local optical activity distributed symmetrically with respect to structural centers. In contrast, achiral chain structures have even inflection points and showed antisymmetric distribution. In the near‐field CD images of fabricated chain nanostructures, the symmetric and antisymmetric distributions of local CD were observed for chiral and achiral chain structures, respectively, consistent with the simulated results. The handedness of the local optical activity was found to be determined by the handedness of the inflection point, for the fabricated chain structures having two or more inflection points. The local optical activity was thus governed primarily by the local chirality of the inflection points for the gold chain structures. The total effect of all the inflection points in the chain structure is considered to be a predominant factor that determines the macroscopic optical activity. Chirality 28:540–544, 2016. © 2016 Wiley Periodicals, Inc.     

Effect of the chelation of metal cation on the antioxidant activity of chondroitin sulfates

The antioxidant potencies of chondroitin sulfates (CSs) from shark cartilage, salmon cartilage, bovine trachea, and porcine intestinal mucosa were compared by three representative methods for the measurement of the antioxidant activity; DPPH radical scavenging activity, superoxide radical scavenging activity, and hydroxyl radical scavenging activity. CSs from salmon cartilage and bovine trachea showed higher potency in comparison with CSs from shark cartilage and porcine intestinal mucosa. Next, CS from salmon cartilage chelating with Ca²⁺, Mg²⁺, Mn²⁺, or Zn²⁺ were prepared, and their antioxidant potencies were compared. CS chelating with Ca²⁺ or Mg²⁺ ions showed rather decreased DPPH radical scavenging activity in comparison with CS of H⁺ form. In contrast, CS chelating with Ca²⁺ or Mg²⁺ ion showed remarkably enhanced superoxide radical scavenging activity than CS of H⁺ or Na⁺ form. Moreover, CS chelating with divalent metal ions, Ca²⁺, Mg²⁺, Mn²⁺, or Zn²⁺, showed noticeably higher hydroxyl radical scavenging activity than CS of H⁺ or Na⁺ form. The present results revealed that the scavenging activities of, at least, superoxide radical and hydroxyl radical were enhanced by the chelation with divalent metal ions.     

Biochemical characteristics and antioxidant activity of crude and purified sulfated polysaccharides from Gracilaria fisheri

Sulfated polysaccharides (SPs) from Gracilaria fisheri of Thailand, which were extracted in low-temperature (25 °C) water showed the highest content of phenolic compounds compared with those extracted at high temperature (55 °C). Crude SP antioxidant activity was evaluated by measuring the DPPH free radical scavenging effect which is directly related to the level of phenolic compounds. The sulfate content, total sugar, and SPs yield were also directly related to the extraction temperature. All extracts contained galactose as a major monosaccharide. High antioxidant activity of crude SP, positively correlated with the phenolic compound contents (R² = 0.996) contributed by the existence of sulfate groups and phenolic compounds. In purified SP, F1 fraction exhibited strong radical scavenging ability, but it was not significantly different compared to crude SP extracted at 25 °C. This indicated that the appropriate density and distribution of sulfate groups in the SP extract showed the best antioxidant activity.     

Anti-bacterial and anti-inflammatory effects of ethanol extract from Houttuynia cordata poultice

Houttuynia cordata (HC) has been commonly used as many traditional remedies in local areas of Japan. Although many pharmacological activities of HC have been reported, the mechanism underlying the effect of HC remains unknown. We conducted the interview survey in Japan to verify how HC was actually used. The interview survey revealed that HC poultice (HCP) prepared from smothering fresh leaves of HC was most frequently used for the treatment of purulent skin diseases including furuncle and carbuncle with high effectiveness. Ethanol extract of HCP (eHCP) showed anti-bacterial effects against methicillin-resistant Staphylococcus aureus (MRSA), and showed an anti-biofilm activity against MRSA. eHCP showed dose-dependent inhibition of S. aureus lipoteichoic acid (LTA)-induced interleukin-8 and CCL20 production in human keratinocyte without any cytotoxicity. These results suggest that HCP is effective for skin abscess and its underlying mechanism might be the complicated multiple activities for both bacteria and host cells.     

Characterization of the reaction of decoupling ubiquinone with bovine mitochondrial respiratory complex I

We previously produced the unique ubiquinone QT (“decoupling” quinone), the catalytic reduction of which in NADH-quinone oxidoreduction with bovine heart mitochondrial NADH-ubiquinone oxidoreductase (complex I) is completely decoupled from proton translocation across the membrane domain. This feature is markedly distinct from those of typical short-chain quinones such as ubiquinone-1. To further characterize the features of the QT reaction with complex I, we herein synthesized three QT analogs, QT2–QT4, and characterized their electron transfer reactions. We found that all aspects of electron transfer (e.g. electron-accepting activity and membrane potential formation) vary significantly among these analogs. The features of QT2 as decoupling quinone were slightly superior to those of original QT. Based on these results, we conclude that the bound positions of QTs within the quinone binding cavity susceptibly change depending on their side-chain structures, and the positions, in turn, govern the behavior of QTs as electron acceptors.     

Circadian rhythm in locomotor activity in the burrower crayfish Procambarus acanthophorus (Villalobos 1948)

Crayfish Procambarus acanthophorus is a burrower that spends long periods building deep tunnels to reach the water table during the dry season; thus, its survival entertains a close ecological relationship with the sediment. The aim of this work was to determine whether the properties of the circadian rhythm of locomotor activity could be modified by the sediment availability. Experiments were conducted in both aquaria filled with sediment or filled with water, under cycles of Bright and Dim Illumination (BI:DI, 12:12 h) or under continuous DI:DI. The rhythm of locomotor activity was entrained with the photoperiod in aquaria with sediment or water; however, statistical differences between conditions were obtained comparing the period and the level of activity under free-running. These data suggest that the substrate’s sensorial perception could be encoded as a significant ecological parameter that exerts influence in the physiological mechanisms that control the temporal order in P. acanthophorus.     

番茄‘Ls-89’和‘坂砧2号’幼苗活性氧代谢对南方根结线虫侵染的反应

为了解番茄(Lycopersicon esculentum)砧木幼苗活性氧代谢与抗南方根结线虫(Meloidogyne incognita)的关系,以高感品种‘Ls-89’与高抗品种‘坂砧2号’为材料,采用盆栽人工接种法,研究了南方根结线虫侵染对番茄砧木幼苗活性氧代谢的影响。结果表明,未接种南方根结线虫的番茄砧木幼苗,其根系与叶片活性氧水平及相关酶活性在品种间没有显著差异。接种南方根结线虫后,两品种幼苗根系与叶片的O_2·~–生成速率和H_2O_2含量均升高,且‘坂砧2号’显著高于‘Ls-89’,但‘Ls-89’的MDA含量则显著高于‘坂砧2号’。两品种幼苗根系与叶片的SOD活性均在侵染早期降低,以‘坂砧2号’降幅较大;但POD、CAT活性在侵染早期变化不大,至侵染中后期则显著升高。因此,番茄抗性品种砧木幼苗的膜脂抗氧化能力较强,活性氧水平较高,且SOD活性对南方根结线虫侵染敏感。