SASDGB6: Resistance to inhibitors of cholinesterase 8 homolog A (Ric8A) mi...

基本情報

• 登録情報: データベース: SASBDB / ID: SASDGB6

試料

Resistance to inhibitors of cholinesterase 8 homolog A (Ric8A) miniGi complex

• Resistance to inhibitors of cholinesterase 8 homolog A (protein), Ric8a, Bos taurus
• miniGi (protein), synthetic construct

機能・相同性

分子機能:

G-protein alpha-subunit binding / guanyl-nucleotide exchange factor activity / G protein-coupled receptor signaling pathway / plasma membrane / cytoplasm

ドメイン・相同性:

Synembryn / Guanine nucleotide exchange factor, Ric8 / Guanine nucleotide exchange factor synembryn / Armadillo-like helical / Armadillo-type fold

構成要素:

Synembryn-A

• 生物種: Bos taurus (ウシ); synthetic construct (人工物)
• 引用: ジャーナル: J Biol Chem / 年: 2019; タイトル: Large-scale conformational rearrangement of the α5-helix of Gα subunits in complex with the guanine nucleotide exchange factor Ric8A.; 著者: Dhiraj Srivastava / Nikolai O Artemyev / ; 要旨: Resistance to inhibitors of cholinesterase 8A (Ric8A) protein is an important G protein-coupled receptor (GPCR)-independent regulator of G protein α-subunits (Gα), acting as a guanine nucleotide exchange factor (GEF) and a chaperone. Insights into the complex between Ric8A and Gα hold the key to understanding the mechanisms underlying noncanonical activation of G-protein signaling as well as the folding of nascent Gα proteins. Here, we examined the structure of the complex of Ric8A with minimized Gα (miniGα) in solution by small-angle X-ray scattering (SAXS) and exploited the scattering profile in modeling of the Ric8A/miniGα complex by steered molecular dynamics (SMD) simulations. A small set of models of the complex featured minimal clash scores, excellent agreement with the experimental SAXS data, and a large-scale rearrangement of the signal-transducing α5-helix of Gα away from its β-sheet core. The resulting interface involved the Gα α5-helix bound to the concave surface of Ric8A and the Gα β-sheet that wraps around the C-terminal part of the Ric8A armadillo domain, leading to a severe disruption of the GDP-binding site. Further modeling of the flexible C-terminal tail of Ric8A indicated that it interacts with the effector surface of Gα. This smaller interface may enable the Ric8A-bound Gα to interact with GTP. The two-interface interaction with Gα described here distinguishes Ric8A from GPCRs and non-GPCR regulators of G-protein signaling.

登録者

• Dhiraj Srivastava (University of Iowa, Iowa, USA)

モデル

• モデル #3814: ; タイプ: atomic / カイ2乗値: 1.637; Omokage検索でこの集合体の類似形状データを探す (詳細)
• モデル #3813: ; タイプ: atomic / ソフトウェア: (Rosetta) / カイ2乗値: 1.882; 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: Resistance to inhibitors of cholinesterase 8 homolog A (Ric8A) miniGi complex; 試料濃度: 10 mg/ml / Entity id: 1575 / 1912
• バッファ: 名称: 20 mM Tris, 150 mM KCl, 5 % glycerol, 1 mM TCEP / pH: 8

要素 #1575

名称: Ric8a / タイプ: protein
記述: Resistance to inhibitors of cholinesterase 8 homolog A
分子量: 55.643 / 分子数: 1 / 由来: Bos taurus / 参照: UniProt: Q5E9J8
配列:

GHMADPRAVA DALETGEEDV VMEALRAYNR ENSQSFTFDD AQQEDRKRLA KLLVSVLEQG LPPSRRVIWL QSIRILSRDR SCLDSFTSRR SLQALACYAG ISASQGSVPE PLNMDVVLES LKCLCNLVLS SPVAQALAAE AGLVVRLAER VGLCRQSSFP HDVQFFDLRL LFLLTALRTD VRQQLFQELQ GVRLLTRALE LTLGMTEGER HPELLPPQET ERAMEILKVL FNITFDSIKR EVDEEDAALY RHLGTLLRHC VMLAAAGDRT EELHGHAVNL LGNLPVKCLD VLLTLEPHEG SLEFLGVNMD VIRVLLSFME KRLHQTHRLK ESVAPVLSVL TECARMHRPA RKFLKAQVLP PLRDVRTRPE VGELLRNKLV RLMTHLDTDV KRVAAEFLFV LCSESVPRFI KYTGYGNAAG LLAARGLMAG GRPEGQYSED EDTDTDEYKE AKASINPVTG RVEEKPPNPM EGMTEEQKEH EAMKLVNMFD KLSRH

要素 #1912

タイプ: protein / 記述: miniGi / 分子量: 24.54 / 分子数: 1 / 由来: synthetic construct
配列:

AMEKAAREVK LLLLGADNSG KSTIVKQMKI IHEAGEYMPM ERVKTTGIVE THFTFKDLHF KMFDVGGQRS ERKKWIHCFE DVAAIIFCVD LSDYEEMNRM HESMKLFDSI CNNKWFTDTS IILFLNKKDL FEEKIKKSPL TICYQEYAGS NTYEEAAAYI QCQFEDLNKR KDTKEIYTHF TCATDTKNVQ FVFDAVTDVI IKNNLKDCGL F

実験情報

• ビーム: 設備名称: Advanced Photon Source (APS), Argonne National Laboratory BioCAT 18ID; 地域: Lemont, IL / 国: USA / 線源: X-ray synchrotron
• 検出器: 名称: Pilatus3 X 1M / Pixsize x: 0.172 mm

スキャン

タイトル: Resistance to inhibitors of cholinesterase 8 homolog A (Ric8A) miniGi complex
測定日: 2018年10月27日 / 保管温度: 4 °C / 照射時間: 0.5 sec. / 単位: 1/A /

	Min	Max
Q	0.0039	0.3813

距離分布関数 P(R)

ソフトウェア P(R): GNOM 4.6 / ポイント数: 420 /

	Min	Max
Q	0.004209	0.3813
P(R) point	1	420
R	0	107

結果

Experimental MW: 83.6 kDa / カーブのタイプ: sec
コメント: Purified Ric8a1-492 was mixed with excess of purified miniGi and the complex was purified by size-exclusion chromatography (SEC). The complex fractions were pooled together, concentrated and additional SEC-SAXS data was collected as described above. X-ray wavelength = UNKNOWN.

	P(R)	P(R) error	Guinier	Guinier error
前方散乱 I0	0.00261	1.93E-6	0.0026	2.65E-6
慣性半径, Rg	3.28 nm	0.003	3.24 nm	0.006

	Min	Max
D	-	10.7
Guinier point	13	118