SASDBY5: C-terminal fragment (509-716) of the Methoprene-tolerant protein ...

Basic information

• Entry: Database: SASBDB / ID: SASDBY5

Sample

C-terminal fragment (509-716) of the Methoprene-tolerant protein from Drosophila melanogaster

• FI10506p (protein), Met, Drosophila melanogaster

Function / homology

Function:

aryl hydrocarbon receptor complex / negative regulation of programmed cell death / regulation of developmental process / transcription factor binding / nuclear receptor activity / DNA-binding transcription factor activity, RNA polymerase II-specific / protein heterodimerization activity / RNA polymerase II cis-regulatory region sequence-specific DNA binding / regulation of transcription by RNA polymerase II / positive regulation of transcription by RNA polymerase II / nucleus

Domain/homology:

Helix-loop-helix DNA-binding domain / Helix-loop-helix DNA-binding domain superfamily / helix loop helix domain / Myc-type, basic helix-loop-helix (bHLH) domain / Myc-type, basic helix-loop-helix (bHLH) domain profile. / PAS domain / PAS repeat profile. / PAS domain / PAS domain superfamily

Component:

FI10506p

• Biological species: Drosophila melanogaster (fruit fly)
• Citation: Journal: PLoS One / Year: 2016; Title: Intrinsic Disorder of the C-Terminal Domain of Drosophila Methoprene-Tolerant Protein.; Authors: Marta Kolonko / Katarzyna Ożga / Rafał Hołubowicz / Michał Taube / Maciej Kozak / Andrzej Ożyhar / Beata Greb-Markiewicz / ; Abstract: Methoprene tolerant protein (Met) has recently been confirmed as the long-sought juvenile hormone (JH) receptor. This protein plays a significant role in the cross-talk of the 20-hydroxyecdysone (20E) and JH signalling pathways, which are important for control of insect development and maturation. Met belongs to the basic helix-loop-helix/Per-Arnt-Sim (bHLH-PAS) family of transcription factors. In these proteins, bHLH domains are typically responsible for DNA binding and dimerization, whereas the PAS domains are crucial for the choice of dimerization partner and the specificity of target gene activation. The C-terminal region is usually responsible for the regulation of protein complex activity. The sequence of the Met C-terminal region (MetC) is not homologous to any sequence deposited in the Protein Data Bank (PDB) and has not been structurally characterized to date. In this study, we show that the MetC exhibits properties typical for an intrinsically disordered protein (IDP). The final averaged structure obtained with small angle X-ray scattering (SAXS) experiments indicates that intrinsically disordered MetC exists in an extended conformation. This extended shape and the long unfolded regions characterise proteins with high flexibility and dynamics. Therefore, we suggest that the multiplicity of conformations adopted by the disordered MetC is crucial for its activity as a biological switch modulating the cross-talk of different signalling pathways in insects.

Contact author

• Michal Taube (AMU, Adam Mickiewicz University in Poznań, Poznań, Poland)

Models

• Model #560: ; Type: dummy / Software: EOM / Radius of dummy atoms: 1.90 A / Chi-square value: 0.962361; Search similar-shape structures of this assembly by Omokage search (details)
• Model #561: ; Type: dummy / Software: EOM / Radius of dummy atoms: 1.90 A / Chi-square value: 0.962361; Search similar-shape structures of this assembly by Omokage search (details)

Sample

• Sample: Name: C-terminal fragment (509-716) of the Methoprene-tolerant protein from Drosophila melanogaster; Specimen concentration: 2.6 mg/ml
• Buffer: Name: 20mM Tris/HCl 150mM NaCl / pH: 7.5

Entity #386

Name: Met / Type: protein / Description: FI10506p / Formula weight: 23.4 / Num. of mol.: 1 / Source: Drosophila melanogaster / References: UniProt: Q9VYW2
Sequence:

GSGGIEGRHA GRQKVQEMKE KFSTIIKAEM PTQSSSPDLP ASQAPQQLER IVLYLIENLQ KSVDSAETVG GQGMESLMDD GYSSPANTLT LEELAPSPTP ALALVPPAPS SVKSSISKSV SVVNVTAARK FQQEHQKQRE RDREQLKERT NSTQGVIRQL SSCLSEAETA SCILSPASSL SASEAPDTPD PHSNTSPPPS LHTRPSVLHR TLTSTLR

Experimental information

• Beam: Instrument name: PETRA III P12 / City: Hamburg / 国: Germany / Type of source: X-ray synchrotron / Wavelength: 0.124 Å / Dist. spec. to detc.: 3 mm
• Detector: Name: Pilatus 2M

Scan

Title: C-terminal fragment (509-716) of the Met / Measurement date: Jul 21, 2015 / Cell temperature: 10 °C / Exposure time: 0.5 sec. / Number of frames: 20 / Unit: 1/nm /

	Min	Max
Q	0.022	4.8204

Distance distribution function P(R)

Sofotware P(R): GNOM 4.6 / Number of points: 412 /

	Min	Max
Q	0.06036	4.405
P(R) point	13	424
R	0	22

Result

D max: 22 / Type of curve: single_conc
Comments: Modelling of the MetC low resolution structure was performed using Ensemble Optimization Method (EOM), of which selected models are displayed in this entry. The full results obtained from EOM modelling that includes the optimised Rg and size distributions of the MetC ensemble are included in the full entry zip archive. Note: The C-terminal fragment of the MetC protein (residues 509-716) has nine additional amino acids (GSGGIEGRH) at the N-terminus derived from the affinity tag encoded in the pColdTF expression plasmid (remaining after protease cleavage). The molecular weight (from I(0)) was calculated from the SAXS data by scaling the intensities by 0.3 in order to match the high-q scattering intensities obtained from a BSA standard. This procedure was done because estimating the concentration the MetC sample is prone to significant errors as a consequence of the protein's poor extinction coefficient (Abs. 280 nm 1mg/ml = 0.130).

	Experimental	Standard
MW	30.52 kDa	30.52 kDa

	P(R)	Guinier
Forward scattering, I0	397600	38540
Radius of gyration, Rg	5.61 nm	5.1 nm