Back to Search
Start Over
Microtubule assembly-derived by dimerization of TPPP/p25. Evaluation of thermodynamic parameters for multiple equilibrium system from ITC data
- Source :
-
BBA - General Subjects . Jul2012, Vol. 1820 Issue 7, p785-794. 10p. - Publication Year :
- 2012
-
Abstract
- Abstract: Background: The disordered Tubulin Polymerization Promoting Protein/p25 (TPPP/p25) modulates the dynamics and stability of the microtubule system. In this paper the role of dimerization in its microtubule-related functions is established, and an approach is proposed to evaluate thermodynamic constants for multiple equilibrium systems from ITC measurements. Methods: For structural studies size exclusion chromatography, SDS-PAGE, chemical cross-linking, circular dichroism, fluorescence spectroscopy and isothermal titration calorimetry were used; the functional effect was analyzed by tubulin polymerization assay. Numerical simulation of the multiple equilibrium was performed with Mathematica software. Results: The dimerization of TPPP/p25 is promoted by elevation of the protein concentration and by GTP addition. The dimeric form displaying enhanced tubulin polymerization promoting activity is stabilized by disulfide bond or chemical cross-linking. The GTP binding to the dimeric form (Kd-GTP =200μM) is tighter with one order of magnitude than to the monomeric one leading to the enrichment of the dimers. A mathematical model elaborated for the multiple equilibrium of the TPPP/p25-GTP system was validated by fitting the GTP-dependent changes of ellipticity and fluorescence signal in the course of TPPP/p25 titrations. The evaluation of the equilibrium constants rendered it possible to determine the thermodynamic parameters of the association of different TPPP/p25 forms with GTP from ITC measurements. Conclusions/General Significance: The dimerization of TPPP/p25 with favorable physiological functional potency is proposed to play significant role in the fine tuning of TPPP/p25-mediated microtubule assembly; the unfolded monomers might be involved in the formation of pathological inclusions characteristic for Parkinson''s disease and other synucleinopathies. [Copyright &y& Elsevier]
Details
- Language :
- English
- ISSN :
- 03044165
- Volume :
- 1820
- Issue :
- 7
- Database :
- Academic Search Index
- Journal :
- BBA - General Subjects
- Publication Type :
- Academic Journal
- Accession number :
- 76158700
- Full Text :
- https://doi.org/10.1016/j.bbagen.2012.03.011