Intestinal-type alkaline phosphatase (IAP) is expressed at a high concentration in the brush border membrane of intestinal epithelial cells and is known to be a gut mucosal defense factor. In humans, ...a single gene (ALPI) for IAP has been isolated, and its transcription produces two kinds of alternatively spliced mRNAs (aAug10 and bAug10). Recently, we discovered that vitamin D up-regulated the expression of both types of human IAP alternative splicing variants in Caco-2 cells. However, the functional difference of protein encoded by the mRNA variants has remained elusive. In the present study, we aimed to provide further insight into the characterization and structure of IAP isoforms. To analyze the protein translated from the ALPI gene, we constructed two kinds of cDNA expression plasmids (aAug10 and bAug10), and the transfected cells were homogenized and assayed for alkaline phosphatase (ALP) activity. We also designed the homology-modeled 3D structures of the protein encoded by the mRNA variants (ALPI-aAug10 and ALPI-bAug10). The levels of ALP activity of COS-1 cells transfected with the aAug10 plasmid were increased significantly, while cells transfected with the bAug10 plasmid had undetectable ALP activity. The homology-modeled 3D structures revealed that the variant bAug10 lacks the central N-terminal α-helix and residue corresponding to Asp-42 of ALPI-aAug10 near the active site. This is the first report on the characterization and structure of alternatively spliced transcript variants of the human ALPI gene. Further studies on the regulation of aAug10 and/or bAug10 mRNA expression may identify novel physiological functions of IAP.
Abstract The lacunar-spinel chalcogenides exhibit magnetic centers in the form of transition-metal tetrahedra. On the basis of density-functional computations, the electronic ground state of an Mo4 ...13+ tetrahedron has been postulated as single-configuration a 1 2 e 4 t 2 5, where a 1, e, and t 2 are symmetry-adapted linear combinations of single-site Mo t 2g atomic orbitals. Here we unveil the many-body tetramer wave-function: we show that sizable correlations yield a weight of only 62% for the a 1 2 e 4 t 2 5 configuration. While spin–orbit coupling within the peculiar valence orbital manifold is still effective, the expectation value of the spin–orbit operator and the g factors deviate from figures describing nominal t 5 j eff = 1/2 moments. As such, our data documents the dressing of a spin–orbit j eff = 1/2 object with intra-tetramer excitations. Our results on the internal degrees of freedom of these magnetic moments provide a solid theoretical starting point in addressing the intriguing phase transitions observed at low temperatures in these materials.
Maleimide-conjugating closo-dodecaborate sodium form 5c (MID) synthesized by the nucleophilic ring-opening reaction of closo-dodecaborate-1,4-dioxane complex 2 with tetrabutylammonium (TBA) azide was ...found to conjugate to free SH of cysteine and lysine residues in BSA under physiological conditions, forming highly boronated BSA that showed high and selective accumulation in tumor and significant tumor growth inhibition in colon 26 tumor-bearing mice subjected to thermal neutron irradiation.
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Selective purification and chemical labeling of a target protein in a protein mixture were simultaneously achieved on the surface of affinity beads functionalized with ligands, such as ...benzenesulfonamide and methotrexate (MTX), and a ruthenium complex containing 2,2'-bipyridine-4,4'-dicarboxylic acid (dcbpy). Chemical labeling of the target protein with a tyrosine radical trapper (TRT) proceeded on the surface of the beads when the target protein was in close proximity to the ruthenium photocatalyst. Both the protein purification and chemical labeling abilities of the affinity beads functionalized with ruthenium photocatalyst were not compromised after recycling several times. Dihydrofolate reductase (DHFR) endogenously expressed in HeLa cells was detected by chemical labeling with biotin-TRT on the affinity beads with high sensitivity compared to the conventional silver staining method.
Feglymycin is a naturally occurring, anti-HIV and antimicrobial 13-mer peptide that includes highly racemizable 3,5-dihydroxyphenylglycines (Dpgs). Here we describe the total synthesis of feglymycin ...based on a linear/convergent hybrid approach. Our originally developed micro-flow amide bond formation enabled highly racemizable peptide chain elongation based on a linear approach that was previously considered impossible. Our developed approach will enable the practical preparation of biologically active oligopeptides that contain highly racemizable amino acids, which are attractive drug candidates.
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•Micro-flow technology enables highly efficient photo reactions.•Integrated micro-flow reaction enables efficient synthesis of complex compounds.•Integrated, multi-step micro-flow ...synthesis containing photo reactions were reviewed.
Micro-flow photochemical reactions have great advantage over batch photochemical reactions due to its high light-penetration efficiency. Integrated micro-flow reaction enables efficient synthesis of structurally complex compounds from simple starting materials and it can avoid handling of explosive, toxic, unstable, or odorous intermediates. Combination of micro-flow photochemical reactions with integrated micro-flow synthesis enhances their benefits. Here we summarize recently reported integrated multi-step micro-flow synthesis containing various photochemical reactions.
The QuantiFERON®-TB Gold In-Tube test (QFT), an interferon-γ release assay, is used to diagnose Mycobacterium tuberculosis, but its inaccuracy in distinguishing active tuberculosis from latent ...infection is a major concern. There is thus a need for an easy and accurate tool for achieving that goal in daily clinical settings. This study aimed to identify candidate cytokines for specifically differentiating active tuberculosis from latent infection. Our study population consisted of 31 active TB (tuberculosis) patients, 29 LTBI (latent tuberculosis infection) patients and 10 healthy control subjects. We assayed for 27 cytokines in QFT supernatants of both specific antigen-stimulated blood samples (TBAg) and negative-control samples (Nil). We analyzed their specificities and sensitivities by creating receiver operating characteristic (ROC) curves and measuring the area under those curves (AUCs). In TBAg-Nil supernatants, IL-10, IFN-γ, MCP-1 and IL-1RA showed high AUCs of 0.8120, 0.7842, 0.7419 and 0.7375, respectively. Compared with each cytokine alone, combined assay for these top four cytokines showed positive rates in diagnosing active TB, and GDA analysis revealed that MCP-1 and IL-5 are potent in distinguishing active TB from LTBI, with Wilk's lambda = 0.718 (p < 0.001). Furthermore, utilizing the unique characteristic of IL-2 that its TBAg-Nil supernatant levels are higher in LTBI compared to active TB, the difference between IFN-γ and IL-2 showed a large AUC of 0.8910. In summary, besides IFN-γ, IL-2, IL-5, IL-10, IL-1RA and MCP-1 in QFT supernatants may be useful for distinguishing active TB from LTBI. Those cytokines may also help us understand the difference in pathogenesis between active TB and LTBI.
The type I interferon (IFN) response is one of the primary defense systems against various pathogens. Although rubella virus (RuV) infection is known to cause dysfunction of various organs and ...systems, including the central nervous system, little is known about how human neural cells evoke protective immunity against RuV infection, leading to controlling RuV replication. Using cultured human neural cells experimentally infected with RuV RA27/3 strain, we characterized the type I IFN immune response against the virus. RuV infected cultured human neural cell lines and induced IFN-β production, leading to the activation of signal transducer and activator of transcription 1 (STAT1) and the increased expression of IFN-stimulated genes (ISGs). Melanoma-differentiation-associated gene 5 (MDA5), one of the cytoplasmic retinoic acid-inducible gene I (RIG-I)-like receptors, is required for the RuV-triggered IFN-β mRNA induction in U373MG cells. We also showed that upregulation of RuV-triggered ISGs was attenuated by blocking IFN-α/β receptor subunit 2 (IFNAR2) using an IFNAR2-specific neutralizing antibody or by repressing mitochondrial antiviral signaling protein (MAVS) expression using MAVS-targeting short hairpin RNA (shRNA). Furthermore, treating RuV-infected cells with BX-795, a TANK-binding kinase 1 (TBK1)/I kappa B kinase ε (IKKε) inhibitor, robustly reduced STAT1 phosphorylation and expression of ISGs, enhancing viral gene expression and infectious virion production. Overall, our findings suggest that the RuV-triggered type I IFN-mediated antiviral response is essential in controlling RuV gene expression and viral replication in human neural cells.
Targeting less abundant amino acid residues on the protein surface may realize site-selective protein modification of natural proteins. The relative hydrophobicity of tyrosine combined with the π–π ...stacking tendency of the aromatic rings results in generally low accessibility. In this study, site-selective protein modification was achieved by targeting surface-exposed tyrosine residues without using a genetic encoding system. Tyrosine residues were modified with N-methylated luminol derivative under single-electron transfer (SET) reaction conditions. Horseradish peroxidase (HRP)-catalyzed SET and electrochemically activated SET modified surface-exposed tyrosine residues selectively. N-Methylated luminol derivative modified tyrosine residues more efficiently than 4-arylurazole under tyrosine click conditions using HRP and electrochemistry. Tyrosine residues that are evolutionarily exposed only in the complementarity-determining region (CDR) of an antibody were selectively modified by tyrosine click reactions. CDR-modified antibodies were applied to in vivo imaging and antibody–drug conjugated (ADC).
Lung cancer frequently co-exists with idiopathic interstitial pneumonia (IIP), which can be subdivided into idiopathic pulmonary fibrosis (IPF) and IIP other than IPF (other IIP). Although ...chemotherapy in small cell lung cancer (SCLC) patients with IIP may result in the exacerbation of IIP, these patients commonly receive chemotherapy. This study aimed to assess the risks and benefits of chemotherapy in SCLC patients with IIP.
We retrospectively analyzed the medical records of 122 patients with SCLC who received chemotherapy. Patients with secondary interstitial lung disease (ILD) of known etiology were excluded. Eligible patients were divided into two groups: SCLC with and without IIP. The former group was subdivided into those with IPF and other IIP.
Of the 47 (39.2%) SCLC patients with IIP, 20 had IPF and 27 had other IIP. The frequency of chemotherapy-induced ILD development or IIP exacerbation was higher in patients with IPF (40.0%) than in those with other IIP (3.7%) and non-IIP (1.4%). Logistic regression analysis demonstrated that ILD development or IIP exacerbation was independently associated with IPF (P = 0.007). Time to treatment failure (P < 0.001) and overall survival (P = 0.001) were different among the groups., Cox proportional hazard model revealed that IPF was independently associated with time to treatment failure (P = 0,017) and overall survival (P = 0.006). Other IIP had no impact on time to treatment failure or overall survival. Development of ILD or exacerbation of IIP independently reduced time to treatment failure and overall survival.
Comorbid IPF can be an independent, negative prognostic indicator and at high risk of ILD development or IIP exacerbation in SCLC patients. Early diagnosis and intervention for chemotherapy-induced IIP exacerbation will be beneficial for SCLC patients with IPF, who need close monitoring for its onset.
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