SASDBU4: Vaccinia primase D5 protein fragment containing the D5N and helic...

基本情報

• 登録情報: データベース: SASBDB / ID: SASDBU4

試料

Vaccinia primase D5 protein fragment containing the D5N and helicase domain

• Primase D5 protein fragment containing the D5N and helicase domain (protein), Vaccinia D5 323-785, Vaccinia virus

機能・相同性

分子機能:

helicase activity / 加水分解酵素; 酸無水物に作用; 酸無水物に作用・細胞または細胞小器官の運動に関与 / host cell cytoplasm / hydrolase activity / ATP binding

ドメイン・相同性:

DNA primase/nucleoside triphosphatase, C-terminal / Poxvirus D5 protein-like / Bacteriophage/plasmid primase, P4, C-terminal / D5 N terminal like / Helicase, superfamily 3, DNA virus / Superfamily 3 helicase of DNA viruses domain profile. / P-loop containing nucleoside triphosphate hydrolase

構成要素:

Uncoating factor OPG117

• 生物種: Vaccinia virus (ワクチニアウイルス)
• 引用: ジャーナル: J Virol / 年: 2016; タイトル: Domain Organization of Vaccinia Virus Helicase-Primase D5.; 著者: Stephanie Hutin / Wai Li Ling / Adam Round / Gregory Effantin / Stefan Reich / Frédéric Iseni / Nicolas Tarbouriech / Guy Schoehn / Wim Pascal Burmeister / ; 要旨: Poxviridae are viruses with a large linear double-stranded DNA genome coding for up to 250 open reading frames and a fully cytoplasmic replication. The double-stranded DNA genome is covalently circularized at both ends. Similar structures of covalently linked extremities of the linear DNA genome are found in the African swine fever virus (asfarvirus) and in the Phycodnaviridae We are studying the machinery which replicates this peculiar genome structure. From our work with vaccinia virus, we give first insights into the overall structure and function of the essential poxvirus virus helicase-primase D5 and show that the active helicase domain of D5 builds a hexameric ring structure. This hexamer has ATPase and, more generally, nucleoside triphosphatase activities that are indistinguishable from the activities of full-length D5 and that are independent of the nature of the base. In addition, hexameric helicase domains bind tightly to single- and double-stranded DNA. Still, the monomeric D5 helicase construct truncated within the D5N domain leads to a well-defined structure, but it does not have ATPase or DNA-binding activity. This shows that the full D5N domain has to be present for hexamerization. This allowed us to assign a function to the D5N domain which is present not only in D5 but also in other viruses of the nucleocytoplasmic large DNA virus (NCLDV) clade. The primase domain and the helicase domain were structurally analyzed via a combination of small-angle X-ray scattering and, when appropriate, electron microscopy, leading to consistent low-resolution models of the different proteins.; IMPORTANCE: Since the beginning of the 1980s, research on the vaccinia virus replication mechanism has basically stalled due to the absence of structural information. As a result, this important class of pathogens is less well understood than most other viruses. This lack of information concerns in general viruses of the NCLDV clade, which use a superfamily 3 helicase for replication, as do poxviruses. Here we provide for the first time information about the domain structure and DNA-binding activity of D5, the poxvirus helicase-primase. This result not only refines the current model of the poxvirus replication fork but also will lead in the long run to a structural basis for antiviral drug design.

登録者

• Stephanie Hutin

モデル

• モデル #534: ; タイプ: dummy / ソフトウェア: DAMMIF (r3709) / ダミー原子の半径: 3.00 A / カイ2乗値: 1.096209 / P-value: 0.000004; Omokage検索でこの集合体の類似形状データを探す (詳細)

試料

• 試料: 名称: Vaccinia primase D5 protein fragment containing the D5N and helicase domain; 試料濃度: 1.42-2.20
• バッファ: 名称: Tris / 濃度: 20.00 mM / pH: 7 / 組成: 150 mM NaCL, 5% glycerol, 1mM DTT

要素 #342

名称: Vaccinia D5 323-785 / タイプ: protein
記述: Primase D5 protein fragment containing the D5N and helicase domain
分子量: 53.48 / 分子数: 6 / 由来: Vaccinia virus / 参照: UniProt: P21010
配列:

GAMGNKLFNI AQRILDTNSV LLTERGDYIV WINNSWKFNS EEPLITKLIL SIRHQLPKEY SSELLCPRKR KTVEANIRDM LVDSVETDTY PDKLPFKNGV LDLVDGMFYS GDDAKKYTCT VSTGFKFDDT KFVEDSPEME ELMNIINDIQ PLTDENKKNR ELYEKTLSSC LCGATKGCLT FFFGETATGK STTKRLLKSA IGDLFVETGQ TILTDVLDKG PNPFIANMHL KRSVFCSELP DFACSGSKKI RSDNIKKLTE PCVIGRPCFS NKINNRNHAT IIIDTNYKPV FDRIDNALMR RIAVVRFRTH FSQPSGREAA ENNDAYDKVK LLDEGLDGKI QNNRYRFAFL YLLVKWYKKY HVPIMKLYPT PEEIPDFAFY LKIGTLLVSS SVKHIPLMTD LSKKGYILYD NVVTLPLTTF QQKISKYFNS RLFGHDIESF INRHKKFANV SDEYLQYIFI EDISSP

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: 0.93 Å / スペクトロメータ・検出器間距離: 2.43 mm
• 検出器: 名称: Pilatus 1M

スキャン

タイトル: Vaccinia virus primase D5, amino acids 323-785
測定日: 2014年9月26日 / 保管温度: 20 °C / セル温度: 20 °C / 照射時間: 2 sec. / フレーム数: 2000 / 単位: 1/nm /

	Min	Max
Q	0.1547	4.9693

距離分布関数 P(R)

ソフトウェア P(R): GNOM 4.6 / ポイント数: 311 /

	Min	Max
Q	0.1973	1.665
P(R) point	10	320
R	0	14.45

結果

カーブのタイプ: single_conc /

	実験値	Porod
分子量	345 kDa	-
体積	-	570 nm3

	P(R)	P(R) error	Guinier	Guinier error
前方散乱 I0	29	0.04	29.5	0.04
慣性半径, Rg	4.7 nm	0.006	4.8 nm	0.03

	Min	Max
D	-	14.5
Guinier point	2	24