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Fluorescence recovery kinetic analysis of gamma-tubulin binding to the mitotic spindle.

Publication ,  Journal Article
Hallen, MA; Ho, J; Yankel, CD; Endow, SA
Published in: Biophys J
September 15, 2008

Fluorescence recovery after photobleaching has been widely used to study dynamic processes in the cell, but less frequently to analyze binding interactions and extract binding constants. Here we use it to analyze gamma-tubulin binding to the mitotic spindle and centrosomes to determine the role of gamma-tubulin in microtubule nucleation in the spindle. We find rapid gamma-tubulin turnover in mitotic spindles of Drosophila early embryos, characterized by diffusional interactions and weak binding, differing from centrosomes with tight binding interactions. The diffusion coefficient of gamma-tubulin is consistent with a major species existing in the cytoplasm as the less efficiently nucleating gamma-tubulin small complex (gammaTuSC) or gamma-tubulin, rather than gamma-tubulin ring complex (gammaTuRC). The fluorescence recovery kinetics we observe implies that gamma-tubulin functions by binding weakly to spindle microtubules. gamma-Tubulin may interact transiently with the spindle, nucleating microtubules very rapidly, differing from centrosomes, where gamma-tubulin binds tightly to nucleate microtubules.

Duke Scholars

Published In

Biophys J

DOI

EISSN

1542-0086

Publication Date

September 15, 2008

Volume

95

Issue

6

Start / End Page

3048 / 3058

Location

United States

Related Subject Headings

  • Tubulin
  • Time Factors
  • Spindle Apparatus
  • Protein Binding
  • Microtubules
  • Kinetics
  • Fluorescence Recovery After Photobleaching
  • Drosophila
  • Diffusion
  • Centrosome
 

Citation

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Hallen, M. A., Ho, J., Yankel, C. D., & Endow, S. A. (2008). Fluorescence recovery kinetic analysis of gamma-tubulin binding to the mitotic spindle. Biophys J, 95(6), 3048–3058. https://doi.org/10.1529/biophysj.108.134593
Hallen, Mark A., Jianghai Ho, Christine D. Yankel, and Sharyn A. Endow. “Fluorescence recovery kinetic analysis of gamma-tubulin binding to the mitotic spindle.Biophys J 95, no. 6 (September 15, 2008): 3048–58. https://doi.org/10.1529/biophysj.108.134593.
Hallen MA, Ho J, Yankel CD, Endow SA. Fluorescence recovery kinetic analysis of gamma-tubulin binding to the mitotic spindle. Biophys J. 2008 Sep 15;95(6):3048–58.
Hallen, Mark A., et al. “Fluorescence recovery kinetic analysis of gamma-tubulin binding to the mitotic spindle.Biophys J, vol. 95, no. 6, Sept. 2008, pp. 3048–58. Pubmed, doi:10.1529/biophysj.108.134593.
Hallen MA, Ho J, Yankel CD, Endow SA. Fluorescence recovery kinetic analysis of gamma-tubulin binding to the mitotic spindle. Biophys J. 2008 Sep 15;95(6):3048–3058.
Journal cover image

Published In

Biophys J

DOI

EISSN

1542-0086

Publication Date

September 15, 2008

Volume

95

Issue

6

Start / End Page

3048 / 3058

Location

United States

Related Subject Headings

  • Tubulin
  • Time Factors
  • Spindle Apparatus
  • Protein Binding
  • Microtubules
  • Kinetics
  • Fluorescence Recovery After Photobleaching
  • Drosophila
  • Diffusion
  • Centrosome