Structural Basis for the 14-3-3 Protein-Dependent Inhibition of
Phosducin Function

J 2017

Structural Basis for the 14-3-3 Protein-Dependent Inhibition of Phosducin Function

KACIROVA, M., Jiří NOVÁČEK, P. MAN, V. OBSILOVA, T. OBSIL et. al.

Basic information

Original name

Structural Basis for the 14-3-3 Protein-Dependent Inhibition of Phosducin Function

Authors

KACIROVA, M. (203 Czech Republic), Jiří NOVÁČEK (203 Czech Republic, guarantor, belonging to the institution), P. MAN (203 Czech Republic), V. OBSILOVA (203 Czech Republic) and T. OBSIL (203 Czech Republic)

Edition

Biophysical Journal, Bethesda, USA, Biophysical Society, 2017, 0006-3495

Other information

Language

English

Type of outcome

Článek v odborném periodiku

Field of Study

10610 Biophysics

Country of publisher

United States of America

Confidentiality degree

není předmětem státního či obchodního tajemství

References:

URL

Impact factor

Impact factor: 3.495

RIV identification code

RIV/00216224:14740/17:00100398

Organization unit

Central European Institute of Technology

DOI

http://dx.doi.org/10.1016/j.bpj.2017.02.036

UT WoS

000398956000007

Keywords in English

EXCHANGE-MASS-SPECTROMETRY; X-RAY-SCATTERING; INTRINSICALLY DISORDERED PROTEINS; SMALL-ANGLE SCATTERING; TRANSDUCIN BETA-GAMMA; LIGAND-BINDING; BIOLOGICAL MACROMOLECULES; PHOSPHORYLATION; REGULATOR; COMPLEX

Abstract

V originále

Phosducin (Pdc) is a conserved phosphoprotein that, when unphosphorylated, binds with high affinity to the complex of bg-subunits of G protein transducin (G(t beta gamma)). The ability of Pdc to bind to G(t beta gamma) is inhibited through its phosphorylation at S54 andS73 within the N-terminaldomain (Pdc-ND) followed by association with the scaffolding protein 14-3-3. However, the molecular basis for the 14-3-3-dependent inhibition of Pdc binding to G(t beta gamma) is unclear. By using small-angle x-ray scattering, high-resolution NMR spectroscopy, and limited proteolysis coupled with mass spectrometry, we show that phosphorylated Pdc and 14-3-3 forma complex in which the Pdc-ND region 45-80, which forms a part of Pdc's G(t beta gamma) binding surface and contains both phosphorylation sites, is restrained within the central channel of the 14-3-3 dimer, with both 14-3-3 binding motifs simultaneously participating in protein association. The N-terminal part of Pdc-NDis likely located outside the central channel of the 14-3-3 dimer, but Pdc residues 20-30, which are also involved in G(t beta gamma) binding, are positioned close to the surface of the 14-3-3 dimer. The C-terminal domain of Pdc is located outside the central channel and its structure is unaffected by the complex formation. These results indicate that the 14-3-3 protein-mediated inhibition of Pdc binding to Gtbg is based on steric occlusion of Pdc's G(t beta gamma) binding surface.

Links

LM2015043, research and development project

Name: Česká infrastruktura pro integrativní strukturní biologii (Acronym: CIISB)

Investor: Ministry of Education, Youth and Sports of the CR

Citovat

KACIROVA, M., Jiří NOVÁČEK, P. MAN, V. OBSILOVA and T. OBSIL. Structural Basis for the 14-3-3 Protein-Dependent Inhibition of Phosducin Function. Biophysical Journal. Bethesda, USA: Biophysical Society, 2017, vol. 112, No 7, p. 1339-1349. ISSN 0006-3495. Available from: https://dx.doi.org/10.1016/j.bpj.2017.02.036.

@article{1411420,
   author = {Kacirova, M. and Nováček, Jiří and Man, P. and Obsilova, V. and Obsil, T.},
   article_location = {Bethesda, USA},
   article_number = {7},
   doi = {http://dx.doi.org/10.1016/j.bpj.2017.02.036},
   keywords = {EXCHANGE-MASS-SPECTROMETRY; X-RAY-SCATTERING; INTRINSICALLY DISORDERED PROTEINS; SMALL-ANGLE SCATTERING; TRANSDUCIN BETA-GAMMA; LIGAND-BINDING; BIOLOGICAL MACROMOLECULES; PHOSPHORYLATION; REGULATOR; COMPLEX},
   language = {eng},
   issn = {0006-3495},
   journal = {Biophysical Journal},
   title = {Structural Basis for the 14-3-3 Protein-Dependent Inhibition of Phosducin Function},
   url = {https://www.sciencedirect.com/science/article/pii/S0006349517302515?via%3Dihub},
   volume = {112},
   year = {2017}
}

TY  - JOUR
ID  - 1411420
AU  - Kacirova, M. - Nováček, Jiří - Man, P. - Obsilova, V. - Obsil, T.
PY  - 2017
TI  - Structural Basis for the 14-3-3 Protein-Dependent Inhibition of Phosducin Function
JF  - Biophysical Journal
VL  - 112
IS  - 7
SP  - 1339-1349
EP  - 1339-1349
PB  - Biophysical Society
SN  - 00063495
KW  - EXCHANGE-MASS-SPECTROMETRY
KW  - X-RAY-SCATTERING
KW  - INTRINSICALLY DISORDERED PROTEINS
KW  - SMALL-ANGLE SCATTERING
KW  - TRANSDUCIN BETA-GAMMA
KW  - LIGAND-BINDING
KW  - BIOLOGICAL MACROMOLECULES
KW  - PHOSPHORYLATION
KW  - REGULATOR
KW  - COMPLEX
UR  - https://www.sciencedirect.com/science/article/pii/S0006349517302515?via%3Dihub
N2  - Phosducin (Pdc) is a conserved phosphoprotein that, when unphosphorylated, binds with high affinity to the complex of bg-subunits of G protein transducin (G(t beta gamma)). The ability of Pdc to bind to G(t beta gamma) is inhibited through its phosphorylation at S54 andS73 within the N-terminaldomain (Pdc-ND) followed by association with the scaffolding protein 14-3-3. However, the molecular basis for the 14-3-3-dependent inhibition of Pdc binding to G(t beta gamma) is unclear. By using small-angle x-ray scattering, high-resolution NMR spectroscopy, and limited proteolysis coupled with mass spectrometry, we show that phosphorylated Pdc and 14-3-3 forma complex in which the Pdc-ND region 45-80, which forms a part of Pdc's G(t beta gamma) binding surface and contains both phosphorylation sites, is restrained within the central channel of the 14-3-3 dimer, with both 14-3-3 binding motifs simultaneously participating in protein association. The N-terminal part of Pdc-NDis likely located outside the central channel of the 14-3-3 dimer, but Pdc residues 20-30, which are also involved in G(t beta gamma) binding, are positioned close to the surface of the 14-3-3 dimer. The C-terminal domain of Pdc is located outside the central channel and its structure is unaffected by the complex formation. These results indicate that the 14-3-3 protein-mediated inhibition of Pdc binding to Gtbg is based on steric occlusion of Pdc's G(t beta gamma) binding surface.
ER  -

KACIROVA, M., Jiří NOVÁČEK, P. MAN, V. OBSILOVA and T. OBSIL. Structural Basis for the 14-3-3 Protein-Dependent Inhibition of Phosducin Function. \textit{Biophysical Journal}. Bethesda, USA: Biophysical Society, 2017, vol.~112, No~7, p.~1339-1349. ISSN~0006-3495. Available from: https://dx.doi.org/10.1016/j.bpj.2017.02.036.

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