1H-NMR investigation of the interaction between RNase T1 and a novel substrate analog, 2′-deoxy-2′-fluoroguanylyl-(3′–5′)uridine

AbstractThe interaction between RNase T1 and a non-hydrolysable substrate analog, 2′-deoxy-2′-fluoroguanylyl-(3′–5′)uridine (GfpU), was investigated using 1H-NMR spectroscopy. In the complex, the Gfp portion takes the syn form around the glycosidic bond and the 3′-endo form for the ribose moiety, similar to those found in 3′-GMP and 2′-deoxy-2′-fluoroguanosine 3′-monophosphate (Gfp). However, in contrast to the cases of these two inhibitors, the complex formation with GfpU at pH 6.0 was found to shift the His-40 C2 proton resonance of RNase T1 to high field as much as 1 ppm. At pH 6.0, this histidine residue appears to be unprotonated in the complex, but is protonated in the free enzyme (pKa of His-40 being 7.9). His-40, rather than Glu-58, is probably involved in the catalytic mechanism as a Lewis base, supporting the recent results from site-directed mutagenesis

Enhanced AKT Phosphorylation of Circulating B Cells in Patients With Activated PI3Kδ Syndrome

Activated PI3Kδ syndrome (APDS) is a primary immunodeficiency characterized by recurrent respiratory tract infections, lymphoproliferation, and defective IgG production. Heterozygous mutations in PIK3CD, PIK3R1, or PTEN, which are related to the hyperactive phosphoinositide 3-kinase (PI3K) signaling, were recently presented to cause APDS1 or APDS2 (APDSs), or APDS-like (APDS-L) disorder. In this study, we examined the AKT phosphorylation of peripheral blood lymphocytes and monocytes in patients with APDSs and APDS-L by using flow cytometry. CD19+ B cells of peripheral blood in APDS2 patients showed the enhanced phosphorylation of AKT at Ser473 (pAKT) without any specific stimulation. The enhanced pAKT in CD19+ B cells was normalized by the addition of a p110δ inhibitor. In contrast, CD3+ T cells and CD14+ monocytes did not show the enhanced pAKT in the absence of stimulation. These findings were similarly observed in patients with APDS1 and APDS-L. Among CD19+ B cells, enhanced pAKT was prominently detected in CD10+ immature B cells compared with CD10− mature B cells. Enhanced pAKT was not observed in B cells of healthy controls, patients with common variable immunodeficiency, and hyper IgM syndrome due to CD40L deficiency. These results suggest that the enhanced pAKT in circulating B cells may be useful for the discrimination of APDS1, APDS2, and APDS-L from other antibody deficiencies.The Supplementary Material for this article can be found online at https://www.frontiersin.org/articles/10.3389/fimmu.2018.00568/full#supplementary-material.This study was supported in part by Grants-in-Aid for Scientific Research from the Japan Society for the Promotion of Science (16H05355 and 16K15528 to SO, 17H04233 to SN), the Ministry of Health, Labour and Welfare, Japan (17933299 to SN), and Practical Research Project for Rare/Intractable Diseases from Japan Agency for Medical Research and Development, AMED

ジュギョウ ヲ コエタ PBL ノ テンカイ コロナカ ニ オケル ジュウガイ プロジェクト ニ カンスル ジッセン ホウコク

近年、野生動物によって引き起こされる獣害が日本各地で大きな社会問題となっている。筆者らは、2019年度に大阪大学で開講されたProject-Based Learning(PBL)1)型授業「フィールド・プロジェクト2)」で、獣害が深刻な滋賀県高島市のX地区周辺で課題解決に取り組んだ。授業は解決策の提案をもって終了したが、提案の改善を重ねて実装に繋げるという過程で実践力を身につけたいという思いから、授業担当教員および地域住民（課題提供者）の協力を得て、PBL形式での学びを継続することにした。新型コロナウイルス感染症拡大によって計画の変更を余儀なくされることもあったが、状況に適応して実現可能なことに着手し、活動を展開した。本活動はPBLを授業だけに留めず、授業終了後も学習目標を掲げ、課題解決に向けて実装に取り組もうとした点で、PBLの継続性に示唆を与え得る事例である。In recent years, agricultural damage caused by wild animals has become a major social problem in various parts of Japan. The authors have tackled the beast damage problem around a district of Takashima City, Shiga Prefecture. This activity started in a project-based-learning (PBL) class coursework called "Field Project" at Osaka University in 2019. The authors made some proposals by the end of the class, but had a desire to acquire practical skills in the process of improving and implementing the pr oposals. So, we decided to continue PBL-style learning in cooperation with a professor and the community residents. However, the pandemic of COVID-19 made us change our plan and there was some limitation to what the authors could do. This activity was beyo nd a class coursework, but we set learning goals and tried to implement our ideas. Therefore, this can give some suggestions to the continuity of PBL