Multistep conformational changes leading to the gate opening of light-driven sodium pump rhodopsin.

Membrane transport proteins require a gating mechanism that opens and closes the substrate transport pathway to carry out unidirectional transport. The "gating" involves large conformational changes and is achieved via multistep reactions. However, these elementary steps have not been clarified for most transporters due to the difficulty of detecting the individual steps. Here, we propose these steps for the gate opening of the bacterial Na+ pump rhodopsin, which outwardly pumps Na+ upon illumination. We herein solved an asymmetric dimer structure of Na+ pump rhodopsin from the bacterium Indibacter alkaliphilus. In one protomer, the Arg108 sidechain is oriented toward the protein center and appears to block a Na+ release pathway to the extracellular (EC) medium. In the other protomer, however, this sidechain swings to the EC side and then opens the release pathway. Assuming that the latter protomer mimics the Na+-releasing intermediate, we examined the mechanism for the swing motion of the Arg108 sidechain. On the EC surface of the first protomer, there is a characteristic cluster consisting of Glu10, Glu159, and Arg242 residues connecting three helices. In contrast, this cluster is disrupted in the second protomer. Our experimental results suggested that this disruption is a key process. The cluster disruption induces the outward movement of the Glu159-Arg242 pair and simultaneously rotates the seventh transmembrane helix. This rotation resultantly opens a space for the swing motion of the Arg108 sidechain. Thus, cluster disruption might occur during the photoreaction and then trigger sequential conformation changes leading to the gate-open state.

Evaluation of a Novel Immunochromatographic Assay Using Monoclonal Antibodies against the Matrix Protein of Human Metapneumovirus.

BACKGROUND: The aim of this study was to determine the sensitivity and specificity of a novel immunochromatographic (IC) assay (APD1806) using monoclonal antibodies against the matrix (M) protein of human metapneumovirus (hMPV) for detection of hMPV from nasopharyngeal swab samples based on the results of real-time RT-PCR. METHODS: Nasopharyngeal swab samples taken from 189 patients aged 0 - 5 years who were suspected of having respiratory tract infections associated with hMPV were used in this study. The samples were tested both by the IC assay and by real-time RT-PCR for detection of hMPV. RESULTS: The sensitivity and specificity of the IC assay for detection of hMPV were 88.8% (95/107) and 92.7% (76/82), respectively. CONCLUSIONS: The IC assay using monoclonal antibodies against the M protein of hMPV is an accurate and fast assay that is suitable as a diagnostic tool for hMPV infection. The optimal timing of the IC assay is 12 hours or more after the onset of fever due to hMPV infection.

Effects of isoflavone-rich red clover extract on blood glucose level: A randomized, double-blind, placebo-controlled trial.

High blood glucose is associated with increased risk of various diseases. Red clover (RC; Trifolium pratense L.) is an edible legume whose sprout is rich in isoflavones such as formononetin and biochanin A. We examined the effects of RC extract on postprandial and fasting blood glucose level, using a randomized, double-blind, placebo-controlled trial with 36 participants, aged 25 to 64 years, who were randomly assigned to receive either 1.91 g of RC extract (containing 8 mg formononetin and 1.8 mg biochanin A) or placebo. Each participant ingested the assigned test food daily for 8 weeks, and at the oral maltose tolerance test (OMTT). Initially, the two groups did not significantly differ in OMTT results. However, fasting insulin levels at 8 weeks were significantly lower in the RC group (4.76 µIU/ml at Week 0 to 4.01 µIU/ml at Week 8) with a significant interaction (P = 0.046). Subgroup analysis showed that change in blood glucose level (blood glucose ΔC) tended to decrease late in the trial period during OMTT in the ≤50-year-old RC group, as did fasting blood glucose and insulin levels at 8 weeks; hemoglobin A1c was also significantly reduced in this subgroup (5.36% at Week 0 to 5.28% at Week 8) with a significant interaction (P = 0.040). These results suggest that the daily intake of RC could reduce blood glucose, particularly for those ≤50 years old. Formononetin-an α-glucosidase inhibitor-is considered to be the major functional molecule for these effects. Therefore, intake of RC that contains formononetin might help blood glucose control.

Low-temperature Raman spectroscopy of sodium-pump rhodopsin from Indibacter alkaliphilus: insight of Na+ binding for active Na+ transport.

We carried out the low-temperature Raman measurement of a sodium pump rhodopsin from Indibacter alkaliphilus (IaNaR) and examined the primary structural change for the light-driven Na+ pump. We observed that photoexcitation of IaNaR produced the distorted 13-cis retinal chromophore in the presence of Na+, while the structural distortion was significantly relaxed in the absence of Na+. This structural difference of the chromophore with/without Na+ was attributed to the Na+ binding to the protein, which alters the active site. Using the spectral sensitivity to the ion binding, we found that IaNaR had a second Na+ binding site in addition to the one already specified on the extracellular surface. To date, the Na+ binding has not been considered as a prerequisite for Na+ transport. However, this study provides insight that the protein structural change induced by the ion binding involved the formation of an R108-D250 salt bridge, which has critical importance in the active transport of Na+.

Evaluation of a Novel Immunochromatographic Assay for Detection of Human Adenoviruses from Throat Swab Samples Compared with Real-time PCR.

BACKGROUND: The aim of this study was to determine the sensitivity and specificity of a novel immunochromatographic assay (ICA) kit, ALSONIC® Adeno (Alfresa Pharma Co., Osaka, Japan), for the detection of human adenovirus (HAdV) from throat swab samples based on the results of real-time PCR. The incubation time required for the novel assay kit (5 minutes) is shorter than that required for other ICA kits that are available in Japan. METHODS: Throat swab samples were taken from 151 patients aged 6 months to 15 years who were suspected of having respiratory tract infections caused by HAdV. RESULTS: The sensitivity and specificity of the ICA for detection of HAdV were 92.2% (83/90) and 95.1% (58/61), respectively, and the assay showed positive and negative predictive values of 96.5% (83/86) and 89.2% (58/65), respectively. CONCLUSIONS: ALSONIC® Adeno is suitable as a diagnostic tool in the acute phase of HAdV infection.

Clinical effectiveness of four neuraminidase inhibitors (oseltamivir, zanamivir, laninamivir, and peramivir) for children with influenza A and B in the 2014-2015 to 2016-2017 influenza seasons in Japan.

The clinical effectiveness of four neuraminidase inhibitors (NAIs) (oseltamivir, zanamivir, laninamivir, and peramivir) for children aged 0 months to 18 years with influenza A and B were investigated in the 2014-2015 to 2016-2017 influenza seasons in Japan. A total of 1207 patients (747 with influenza A and 460 with influenza B) were enrolled. The Cox proportional-hazards model using all of the patients showed that the duration of fever after administration of the first dose of the NAI was shorter in older patients (hazard ratio = 1.06 per 1 year of age, p < 0.001) and that the duration of fever after administration of the first dose of the NAI was shorter in patients with influenza A infection than in patients with influenza B infection (hazard ratio = 2.21, p < 0.001). A logistic regression model showed that the number of biphasic fever episodes was 2.99-times greater for influenza B-infected patients than for influenza A-infected patients (p < 0.001). The number of biphasic fever episodes in influenza A- or B-infected patients aged 0-4 years was 2.89-times greater than that in patients aged 10-18 years (p = 0.010), and the number of episodes in influenza A- or B-infected patients aged 5-9 years was 2.13-times greater than that in patients aged 10-18 years (p = 0.012).

A case of intraocular yolk sac tumor in a child and its pathogenesis.

While yolk sac tumor is one of the most common malignant germ cell tumors occurring in young children, it is rarely found in extragonadal sites. We report a case of intraocular yolk sac tumor in a 4-year-old boy. The diagnosis was confirmed by histologic examination and by the rapid normalization of serum alpha-fetoprotein level following enucleation. We propose that yolk sac cells can potentially migrate into the eye at 22 days of embryonic life during neural tube formation, when the head and tail of the neuropore open contemporaneously and communicate with the amniotic cavity.