SASDE53: Unlabeled dihydrolipoyllysine-residue succinyltransferase compone...

基本情報

登録情報

データベース: SASBDB / ID: SASDE53

試料

Unlabeled dihydrolipoyllysine-residue succinyltransferase component (BBL) with denaturant

• Dihydrolipoyllysine-residue succinyltransferase component of 2-oxoglutarate dehydrogenase complex (protein), BBL, Escherichia coli (strain K12)

機能・相同性

分子機能:

L-lysine catabolic process to acetyl-CoA via saccharopine / dihydrolipoyllysine-residue succinyltransferase / dihydrolipoyllysine-residue succinyltransferase activity / lipoic acid binding / oxoglutarate dehydrogenase complex / tricarboxylic acid cycle / cytosol / cytoplasm

ドメイン・相同性:

Dihydrolipoamide succinyltransferase / : / Peripheral subunit-binding domain / e3 binding domain / Peripheral subunit-binding (PSBD) domain profile. / E3-binding domain superfamily / 2-oxo acid dehydrogenase, lipoyl-binding site / 2-oxo acid dehydrogenases acyltransferase component lipoyl binding site. / 2-oxoacid dehydrogenase acyltransferase, catalytic domain / 2-oxoacid dehydrogenases acyltransferase (catalytic domain) / Biotin-requiring enzyme / Biotinyl/lipoyl domain profile. / Biotin/lipoyl attachment / Single hybrid motif / Chloramphenicol acetyltransferase-like domain superfamily

構成要素:

Dihydrolipoyllysine-residue succinyltransferase component of 2-oxoglutarate dehydrogenase complex

• 生物種: Escherichia coli (strain K12) (大腸菌)
• 引用: ジャーナル: Proc Natl Acad Sci U S A / 年: 2017; タイトル: Decoupling of size and shape fluctuations in heteropolymeric sequences reconciles discrepancies in SAXS vs. FRET measurements.; 著者: Gustavo Fuertes / Niccolò Banterle / Kiersten M Ruff / Aritra Chowdhury / Davide Mercadante / Christine Koehler / Michael Kachala / Gemma Estrada Girona / Sigrid Milles / Ankur Mishra / Patrick R Onck / Frauke Gräter / Santiago Esteban-Martín / Rohit V Pappu / Dmitri I Svergun / Edward A Lemke / ; 要旨: Unfolded states of proteins and native states of intrinsically disordered proteins (IDPs) populate heterogeneous conformational ensembles in solution. The average sizes of these heterogeneous systems, quantified by the radius of gyration ( ), can be measured by small-angle X-ray scattering (SAXS). Another parameter, the mean dye-to-dye distance ( ) for proteins with fluorescently labeled termini, can be estimated using single-molecule Förster resonance energy transfer (smFRET). A number of studies have reported inconsistencies in inferences drawn from the two sets of measurements for the dimensions of unfolded proteins and IDPs in the absence of chemical denaturants. These differences are typically attributed to the influence of fluorescent labels used in smFRET and to the impact of high concentrations and averaging features of SAXS. By measuring the dimensions of a collection of labeled and unlabeled polypeptides using smFRET and SAXS, we directly assessed the contributions of dyes to the experimental values and For chemically denatured proteins we obtain mutual consistency in our inferences based on and , whereas for IDPs under native conditions, we find substantial deviations. Using computations, we show that discrepant inferences are neither due to methodological shortcomings of specific measurements nor due to artifacts of dyes. Instead, our analysis suggests that chemical heterogeneity in heteropolymeric systems leads to a decoupling between and that is amplified in the absence of denaturants. Therefore, joint assessments of and combined with measurements of polymer shapes should provide a consistent and complete picture of the underlying ensembles.

登録者

• Gustavo Fuertes Vives (Institute of Biotechnology CAS v.v.i.)

モデル

試料

• 試料: 名称: Unlabeled dihydrolipoyllysine-residue succinyltransferase component (BBL) with denaturant; 試料濃度: 2 mg/ml
• バッファ: 名称: PBS, 10 mM DTT, 6 M urea, 0.3 M KCl / pH: 7.4

要素 #1242

名称: BBL / タイプ: protein
記述: Dihydrolipoyllysine-residue succinyltransferase component of 2-oxoglutarate dehydrogenase complex
分子量: 4.348 / 分子数: 1 / 由来: Escherichia coli (strain K12) / 参照: UniProt: P0AFG6
配列:

ACSPAIRRLL AEHNLDASAI KGTGVGGRLT REDVEKHLAU A

実験情報

• ビーム: 設備名称: PETRA III EMBL P12 / 地域: Hamburg / 国: Germany / 線源: X-ray synchrotron / 波長: 0.1 Å / スペクトロメータ・検出器間距離: 3 mm
• 検出器: 名称: Pilatus 2M

スキャン

タイトル: Unlabeled dihydrolipoyllysine-residue succinyltransferase component, BBL, in denaturant (urea 6 M)
測定日: 2013年11月8日 / セル温度: 23 °C / 照射時間: 0.05 sec. / フレーム数: 20 / 単位: 1/nm /

	Min	Max
Q	0.0291	4.4884

距離分布関数 P(R)

ソフトウェア P(R): GNOM 5.0 / ポイント数: 1156 /

	Min	Max
Q	0.155497	3.19777
P(R) point	1	1156
R	0	8.26

結果

カーブのタイプ: single_conc
コメント: The protein contains a penultimate non-canonical amino acid p-acetylphenylalanine (207 Da) that is represented as U (selenocysteine, 168 Da) in the amino acid sequence for the entry. Therefore, the calculated MW from sequence (MW(expected)) must be adjusted accordingly (ca. 40 Da).

	実験値	Porod
分子量	10.4 kDa	-
体積	-	41.1 nm3

	P(R)	P(R) error	Guinier	Guinier error
前方散乱 I0	863	17.6	870	8.7
慣性半径, Rg	2.095 nm	0.11	2.1 nm	0.4

	Min	Max
D	-	8.26
Guinier point	49	231