このアイテムのアクセス数: 107

このアイテムのファイル:
ファイル 記述 サイズフォーマット 
fmicb.2016.00587.pdf2.98 MBAdobe PDF見る/開く
タイトル: Characterization of the deamination coupled with sliding along DNA of anti-HIV factor APOBEC3G on the basis of the pH-dependence of deamination revealed by real-time NMR monitoring
著者: Kamba, Keisuke
Nagata, Takashi  kyouindb  KAKEN_id
Katahira, Masato  kyouindb  KAKEN_id
著者名の別形: 永田, 崇
片平, 正人
キーワード: APOBEC3G
NMR
monitoring
enzyme
HIV-1
発行日: 28-Apr-2016
出版者: Frontiers
誌名: Frontiers in Microbiology
巻: 7
論文番号: 587
抄録: Human APOBEC3G (A3G) is an antiviral factor that inactivates HIV. The C-terminal domain of A3G (A3G-CTD) deaminates cytosines into uracils within single-stranded DNA (ssDNA), which is reverse-transcribed from the viral RNA genome. The deaminase activity of A3G is highly sequence-specific; the third position (underlined) of a triplet cytosine (CCC) hotspot is converted into CCU. A3G deaminates a CCC that is located close to the 5' end of ssDNA more effectively than ones that are less close to the 5' end, so-called 3' → 5' polarity. We had developed an NMR method that can be used to analyze the deamination reaction in real-time. Using this method, we previously showed that 3' → 5' polarity can be explained rationally by A3G-CTD's nonspecific ssDNA-binding and sliding direction-dependent deamination activities. We then demonstrated that the phosphate backbone is important for A3G-CTD to slide on the ssDNA and to exert the 3' → 5' polarity, probably due to an electrostatic intermolecular interaction. In this study, we investigate the pH effects on the structure, deaminase activity, and 3' → 5' polarity of A3G-CTD. Firstly, A3G-CTD was shown to retain the native structure in the pH range of 4.0-10.5 by CD spectroscopy. Next, deamination assaying involving real-time NMR spectroscopy for 10-mer ssDNA containing a single CCC revealed that A3G-CTD's deaminase activity decreases as the pH increases in the range of pH 6.5-12.7. This is explained by destabilization of the complex between A3G-CTD and ssDNA due to the weakened electrostatic interaction with the increase in pH. Finally, deamination assaying for 38-mer ssDNA having two CCC hotspots connected by a long poly-adenine linker showed that A3G-CTD retains the same pH deaminase activity preference toward each CCC as that toward the CCC of the 10-mer DNA. Importantly, the 3' → 5' polarity turned out to increase as the pH decreases in the range of 6.5-8.0. This suggests that A3G-CTD tends to continue sliding without abortion at lower pH, while A3G-CTD tends to dissociate from ssDNA during sliding at higher pH due to the weakened electrostatic interaction.
著作権等: © 2016 Kamba, Nagata and Katahira. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
URI: http://hdl.handle.net/2433/214289
DOI(出版社版): 10.3389/fmicb.2016.00587
出現コレクション:学術雑誌掲載論文等

アイテムの詳細レコードを表示する

Export to RefWorks


出力フォーマット 


このリポジトリに保管されているアイテムはすべて著作権により保護されています。