SASDGR2: Thymine dioxygenase J-containing DNA binding domain (JDBD)

基本情報

登録情報

データベース: SASBDB / ID: SASDGR2

試料

Thymine dioxygenase J-containing DNA binding domain (JDBD)

• J-DNA binding domain (protein), JDBD, Leishmania tarentolae

機能・相同性

分子機能:

thymine dioxygenase / thymine dioxygenase activity / base J metabolic process / DNA binding / metal ion binding / nucleus

ドメイン・相同性:

Thymine dioxygenase JBP1, DNA-binding domain / Thymine dioxygenase JBP1 DNA-binding domain / JBP1, DNA-binding domain superfamily / 2OGFeDO, oxygenase domain / Oxygenase domain of the 2OGFeDO superfamily

構成要素:

Thymine dioxygenase JBP1

• 生物種: Leishmania tarentolae (真核生物)
• 引用: ジャーナル: J Biol Chem / 年: 2019; タイトル: The domain architecture of the protozoan protein J-DNA-binding protein 1 suggests synergy between base J DNA binding and thymidine hydroxylase activity.; 著者: Athanassios Adamopoulos / Tatjana Heidebrecht / Jeroen Roosendaal / Wouter G Touw / Isabelle Q Phan / Jos Beijnen / Anastassis Perrakis / ; 要旨: J-DNA-binding protein 1 (JBP1) contributes to the biosynthesis and maintenance of base J (β-d-glucosyl-hydroxymethyluracil), an epigenetic modification of thymidine (T) confined to pathogenic protozoa such as and JBP1 has two known functional domains: an N-terminal T hydroxylase (TH) homologous to the 5-methylcytosine hydroxylase domain in TET proteins and a J-DNA-binding domain (JDBD) that resides in the middle of JBP1. Here, we show that removing JDBD from JBP1 results in a soluble protein (Δ-JDBD) with the N- and C-terminal regions tightly associated together in a well-ordered structure. We found that this Δ-JDBD domain retains TH activity but displays a 15-fold lower apparent rate of hydroxylation compared with JBP1. Small-angle X-ray scattering (SAXS) experiments on JBP1 and JDBD in the presence or absence of J-DNA and on Δ-JDBD enabled us to generate low-resolution three-dimensional models. We conclude that Δ-JDBD, and not the N-terminal region of JBP1 alone, is a distinct folding unit. Our SAXS-based model supports the notion that binding of JDBD specifically to J-DNA can facilitate T hydroxylation 12-14 bp downstream on the complementary strand of the J-recognition site. We postulate that insertion of the JDBD module into the Δ-JDBD scaffold during evolution provided a mechanism that synergized J recognition and T hydroxylation, ensuring inheritance of base J in specific sequence patterns following DNA replication in kinetoplastid parasites.

登録者

• Nassos Adamopoulos (Netherlands Cancer Institute, Amsterdam, Netherlands)

モデル

試料

• 試料: 名称: Thymine dioxygenase J-containing DNA binding domain (JDBD); 試料濃度: 3.75 mg/ml
• バッファ: 名称: 20 mM HEPES, 200 mM NaCl, 1 mM TCEP / pH: 7.5

要素 #898

名称: JDBD / タイプ: protein / 記述: J-DNA binding domain / 分子量: 21.244 / 分子数: 1 / 由来: Leishmania tarentolae / 参照: UniProt: Q9U6M1
配列:

GPGPLSRLGG FSETNLMVST AVEKKKYLDS EFLLHCISAQ LLDMWKQARA RWLELVGKEW AHMLALNPER KDFLWKNQSE MNSAFFDLCE VGKQVMLGLL GKEVALPKEE QAFWIMYAVH LSAACAEELH MPEVAMSLRK LNVKLKDFNF GGTRYFKDMP PEEKKRRMER KQRIEEARRH GMP

実験情報

• ビーム: 設備名称: ESRF BM29 / 地域: Grenoble / 国: France / 線源: X-ray synchrotron / 波長: 0.1 Å / スペクトロメータ・検出器間距離: 2.867 mm
• 検出器: 名称: Pilatus 1M / タイプ: Dectris / Pixsize x: 172 mm

スキャン

タイトル: Thymine dioxygenase J-containing DNA binding domain (JDBD)
測定日: 2017年2月4日 / セル温度: 4 °C / 照射時間: 1 sec. / フレーム数: 34 / 単位: 1/nm /

	Min	Max
Q	0.129	4.9417

距離分布関数 P(R)

ソフトウェア P(R): GNOM 5.0 / ポイント数: 881 /

	Min	Max
Q	0.218449	4.36247
P(R) point	1	881
R	0	7.1

結果

カーブのタイプ: sec /

	実験値	Porod
分子量	28.3 kDa	24 kDa
体積	-	38 nm3

	P(R)	Guinier	Guinier error
前方散乱 I0	7.097	7.14	0.027
慣性半径, Rg	2.177 nm	2.18 nm	0.03

	Min	Max
D	-	7.1
Guinier point	20	100