SASDCN3: Phosphoenolpyruvate-protein phosphotransferase (Enzyme I)

Basic information

• Entry: Database: SASBDB / ID: SASDCN3

Sample

Phosphoenolpyruvate-protein phosphotransferase

• Phosphoenolpyruvate-protein phosphotransferase (protein), Enzyme I, Escherichia coli

Function / homology

Function:

phosphoenolpyruvate-protein phosphotransferase / phosphoenolpyruvate-protein phosphotransferase activity / phosphoenolpyruvate-dependent sugar phosphotransferase system / kinase activity / metal ion binding / cytoplasm

Domain/homology:

Phosphotransferase system, enzyme I / Phosphotransferase system, enzyme I-like / Phosphotransferase system, enzyme I N-terminal / PtsI, HPr-binding domain superfamily / PEP-utilising enzyme, N-terminal / PEP-utilising enzyme, active site / PEP-utilizing enzymes phosphorylation site signature. / PEP-utilising enzyme, conserved site / PEP-utilizing enzymes signature 2. / PEP-utilising enzyme, C-terminal / PEP-utilising enzyme, PEP-binding domain / PEP-utilising enzyme, mobile domain / Phosphohistidine domain superfamily / PEP-utilising enzyme, mobile domain / Pyruvate kinase-like domain superfamily / Pyruvate/Phosphoenolpyruvate kinase-like domain superfamily

Component:

Phosphoenolpyruvate-protein phosphotransferase

• Biological species: Escherichia coli (E. coli)
• Citation: Journal: J Am Chem Soc / Year: 2010; Title: Solution structure of the 128 kDa enzyme I dimer from Escherichia coli and its 146 kDa complex with HPr using residual dipolar couplings and small- and wide-angle X-ray scattering.; Authors: Charles D Schwieters / Jeong-Yong Suh / Alexander Grishaev / Rodolfo Ghirlando / Yuki Takayama / G Marius Clore / ; Abstract: The solution structures of free Enzyme I (EI, ∼128 kDa, 575 × 2 residues), the first enzyme in the bacterial phosphotransferase system, and its complex with HPr (∼146 kDa) have been solved using novel methodology that makes use of prior structural knowledge (namely, the structures of the dimeric EIC domain and the isolated EIN domain both free and complexed to HPr), combined with residual dipolar coupling (RDC), small- (SAXS) and wide- (WAXS) angle X-ray scattering and small-angle neutron scattering (SANS) data. The calculational strategy employs conjoined rigid body/torsion/Cartesian simulated annealing, and incorporates improvements in calculating and refining against SAXS/WAXS data that take into account complex molecular shapes in the description of the solvent layer resulting in a better representation of the SAXS/WAXS data. The RDC data orient the symmetrically related EIN domains relative to the C(2) symmetry axis of the EIC dimer, while translational, shape, and size information is provided by SAXS/WAXS. The resulting structures are independently validated by SANS. Comparison of the structures of the free EI and the EI-HPr complex with that of the crystal structure of a trapped phosphorylated EI intermediate reveals large (∼70-90°) hinge body rotations of the two subdomains comprising the EIN domain, as well as of the EIN domain relative to the dimeric EIC domain. These large-scale interdomain motions shed light on the structural transitions that accompany the catalytic cycle of EI.

Models

• Model #1248: ; Type: atomic / Software: PDB / Radius of dummy atoms: 1.90 A / Chi-square value: 0.341; Search similar-shape structures of this assembly by Omokage search (details)

Sample

• Sample: Name: Phosphoenolpyruvate-protein phosphotransferase / Specimen concentration: 5 mg/ml
• Buffer: Name: 20mM TRIS buffer, 100 mM NaCl, 10 mM DTT, 4 mM MgCl2, 1 mM EDTA; pH: 7.4 ; Comment: 1 tablet of protease inhibitor cocktail (SigmaFAST S8830) was added

Entity #658

Name: Enzyme I / Type: protein / Description: Phosphoenolpyruvate-protein phosphotransferase / Formula weight: 63.561 / Num. of mol.: 2 / Source: Escherichia coli / References: UniProt: A0A037YGN3
Sequence:

MISGILASPG IAFGKALLLK EDEIVIDRKK ISADQVDQEV ERFLSGRAKA SAQLETIKTK AGETFGEEKE AIFEGHIMLL EDEELEQEII ALIKDKHMTA DAAAHEVIEG QASALEELDD EYLKERAADV RDIGKRLLRN ILGLKIIDLS AIQDEVILVA ADLTPSETAQ LNLKKVLGFI TDAGGRTSHT SIMARSLELP AIVGTGSVTS QVKNDDYLIL DAVNNQVYVN PTNEVIDKMR AVQEQVASEK AELAKLKDLP AITLDGHQVE VCANIGTVRD VEGAERNGAE GVGLYRTEFL FMDRDALPTE EEQFAAYKAV AEACGSQAVI VRTMDIGGDK ELPYMNFPKE ENPFLGWRAI RIAMDRREIL RDQLRAILRA SAFGKLRIMF PMIISVEEVR ALRKEIEIYK QELRDEGKAF DESIEIGVMV ETPAAATIAR HLAKEVDFFS IGTNDLTQYT LAVDRGNDMI SHLYQPMSPS VLNLIKQVID ASHAEGKWTG MCGELAGDER ATLLLLGMGL DEFSMSAISI PRIKKIIRNT NFEDAKVLAE QALAQPTTDE LMTLVNKFIE EKTIC

Experimental information

• Beam: Instrument name: Advanced Photon Source (APS) 12ID-C / City: Argonne, IL / 国: USA / Type of source: X-ray synchrotron / Wavelength: 0.06199 Å / Dist. spec. to detc.: 4 mm
• Detector: Name: Gold CCD

Scan

Title: Phosphoenolpyruvate-protein phosphotransferase / Measurement date: Aug 23, 2010 / Cell temperature: 25 °C / Exposure time: 0.25 sec. / Number of frames: 20 / Unit: 1/A /

	Min	Max
Q	0.0144	0.2208

Distance distribution function P(R)

Sofotware P(R): GNOM 5.0 / Number of points: 164 /

	Min	Max
Q	0.014426	0.1953
P(R) point	1	164
R	0	148.6

Result

Type of curve: single_conc /

	Experimental	Porod
MW	128 kDa	118 kDa
Volume	-	189 nm3

	Guinier	P(R)	P(R) error	Guinier error
Forward scattering, I0	0.0290498	-	-	-
Radius of gyration, Rg	4.1 nm	4.2 nm	0.02	0.06

	Max	Error
D	14.75	0.5