SASDAW6: Iron-sulfur cluster assembly scaffold IscU monomer

Basic information

• Entry: Database: SASBDB / ID: SASDAW6

Sample

Iron-sulfur cluster assembly scaffold IscU monomer

• Iron-sulfur cluster assembly scaffold protein IscU (protein), Escherichia coli

Function / homology

Function:

IscS-IscU complex / L-cysteine catabolic process / iron-sulfur cluster assembly / ferrous iron binding / 2 iron, 2 sulfur cluster binding / 4 iron, 4 sulfur cluster binding / intracellular iron ion homeostasis / copper ion binding / zinc ion binding / identical protein binding / cytosol / cytoplasm

Domain/homology:

ISC system FeS cluster assembly, IscU scaffold / NIF system FeS cluster assembly, NifU, N-terminal / NifU-like N terminal domain

Component:

Iron-sulfur cluster assembly scaffold protein IscU

• Biological species: Escherichia coli (E. coli)
• Citation: Journal: Nat Commun / Year: 2010; Title: Structural bases for the interaction of frataxin with the central components of iron-sulphur cluster assembly.; Authors: Filippo Prischi / Petr V Konarev / Clara Iannuzzi / Chiara Pastore / Salvatore Adinolfi / Stephen R Martin / Dmitri I Svergun / Annalisa Pastore / ; Abstract: Reduced levels of frataxin, an essential protein of as yet unknown function, are responsible for causing the neurodegenerative pathology Friedreich's ataxia. Independent reports have linked frataxin to iron-sulphur cluster assembly through interactions with the two central components of this machinery: desulphurase Nfs1/IscS and the scaffold protein Isu/IscU. In this study, we use a combination of biophysical methods to define the structural bases of the interaction of CyaY (the bacterial orthologue of frataxin) with the IscS/IscU complex. We show that CyaY binds IscS as a monomer in a pocket between the active site and the IscS dimer interface. Recognition does not require iron and occurs through electrostatic interactions of complementary charged residues. Mutations at the complex interface affect the rates of enzymatic cluster formation. CyaY binding strengthens the affinity of the IscS/IscU complex. Our data suggest a new paradigm for understanding the role of frataxin as a regulator of IscS functions.

Contact author

• Petr Konarev (EMBL-Hamburg, European Molecular Biology Laboratory (EMBL) - Hamburg outstation, Notkestraße 85, Geb. 25A, 22607 Hamburg, Deutschland, Germany)

Models

• Model #223: ; Type: atomic / Software: crysol / Radius of dummy atoms: 1.90 A / Symmetry: P1 / Chi-square value: 2.199289; Search similar-shape structures of this assembly by Omokage search (details)
• Model #224: ; Type: dummy / Software: DAMMIF / Radius of dummy atoms: 2.30 A / Symmetry: P1 / Chi-square value: 1.065024; Search similar-shape structures of this assembly by Omokage search (details)

Sample

• Sample: Name: Iron-sulfur cluster assembly scaffold IscU monomer / Specimen concentration: 2.5 mg/ml
• Buffer: Name: Tris-HCl / Concentration: 20.00 mM / pH: 8 / Composition: 150 mM NaCl and 10 mM β-mercaptoethanol

Entity #136

Type: protein
Description: Iron-sulfur cluster assembly scaffold protein IscU
Formula weight: 14.464 / Num. of mol.: 1 / Source: Escherichia coli / References: UniProt: P0ACD4
Sequence:

MAYSEKVIDH YENPRNVGSL DKKDSNVGTG MVGAPACGDV MQLQIKVDDN GIIEDAKFKT YGCGSAIASS SLITEWVKGK SLEEAGAIKN SQIAEELELP PVKVHCSILA EDAIKAAIAD YKAKQGLEHH HHHH

Experimental information

• Beam: Instrument name: DORIS III X33 / City: Hamburg / 国: Germany / Shape: 0.6 / Type of source: X-ray synchrotron / Wavelength: 0.15 Å / Dist. spec. to detc.: 2.7 mm
• Detector: Name: Pilatus 1M-W / Pixsize x: 0.172 mm

Scan

Title: u / Measurement date: Nov 5, 2009 / Storage temperature: 15 °C / Cell temperature: 15 °C / Exposure time: 30 sec. / Number of frames: 4 / Unit: 1/nm /

	Min	Max
Q	0.1521	6.237

Distance distribution function P(R)

Sofotware P(R): GNOM 4.5a / Number of points: 484 /

	Min	Max
Q	0.3055	5.457
P(R) point	85	568
R	0	6.41

Result

Type of curve: single_conc
Comments: Reduced levels of frataxin, an essential protein of as yet unknown function, are responsible for causing the neurodegenerative pathology Friedreich’s ataxia. Independent reports have linked frataxin to iron–sulphur cluster assembly through interactions with the two central components of this machinery: desulphurase Nfs1/IscS and the scaffold protein Isu/IscU. In this study, we use a combination of biophysical methods to define the structural bases of the interaction of CyaY (the bacterial orthologue of frataxin) with the IscS/IscU complex. We show that CyaY binds IscS as a monomer in a pocket between the active site and the IscS dimer interface. Recognition does not require iron and occurs through electrostatic interactions of complementary charged residues. Mutations at the complex interface affect the rates of enzymatic cluster formation. CyaY binding strengthens the affinity of the IscS/IscU complex. Our data suggest a new paradigm for understanding the role of frataxin as a regulator of IscS functions.

	Experimental	Standard
MW	14 kDa	14 kDa

	P(R)	Guinier	Guinier error
Forward scattering, I0	17.2	17.2	-
Radius of gyration, Rg	1.86 nm	1.87 nm	0.006

	Min	Max
D	-	6.4
Guinier point	84	-