Related Experiment Videos
Tubulin exchanges divalent cations at both guanine nucleotide-binding sites
J J Correia1, A H Beth, R C Williams
1Department of Molecular Biology, Vanderbilt University, Nashville, Tennessee 37235.
The Journal of Biological Chemistry
|August 5, 1988
Summary
Tubulin
Area of Science:
- Biochemistry
- Molecular Biology
- Structural Biology
Background:
- Tubulin heterodimers bind GTP/GDP at nucleotide-binding sites.
- Magnesium ions (Mg2+) are crucial for GTP binding at the exchangeable site (E-site).
Purpose of the Study:
- Investigate the role and affinity of divalent cations (Mg2+, Mn2+) at tubulin's nucleotide-binding sites.
- Elucidate the mechanism of metal ion exchange at the nonexchangeable site (N-site).
Main Methods:
- Utilized atomic absorption and Electron Paramagnetic Resonance (EPR) spectroscopy.
- Measured metal ion (Mn2+ for Mg2+) exchange rates and binding affinities.
- Assessed tubulin secondary structure and microtubule formation ability.
Main Results:
- Tubulin with GDP at the E-site has one high-affinity metal-binding site; tubulin with GTP has two.
- Mn2+ exhibits higher affinity than Mg2+ for the E-site.
- N-site metal ion exchange is slow and may involve protein breathing, independent of protein/metal concentration.
- Metal ion exchange does not affect tubulin's structure or microtubule assembly.
Conclusions:
- Divalent cations are integral to GTP binding at both N- and E-sites of tubulin.
- Protein dynamics, potentially "breathing," likely govern N-site metal ion release.
- Findings impact the interpretation of structural studies using paramagnetic ions and microtubule dynamics.