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High affinity DNA-microtubule associated protein interaction.

K A Marx1

  • 1Department of Chemistry, University of Massachusetts, Lowell 01854.

Molecular and Cellular Biochemistry
|July 6, 1992
PubMed
Summary

Microtubule-associated proteins/tau (MAP/tau) bind DNA with conserved sequences. These interactions are sequence-specific and do not require tubulin, indicating a fundamental role for MAP/tau in DNA binding.

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • Genetics

Background:

  • Microtubule-associated proteins/tau (MAP/tau) are known to be involved in various cellular processes.
  • Previous studies have indicated that MAP/tau proteins possess DNA binding activity.
  • The specific nature and conservation of this DNA binding activity remain to be fully elucidated.

Purpose of the Study:

  • To isolate and characterize the DNA binding activity of MAP/tau proteins from chick brain microtubule preparations.
  • To determine the affinity and specificity of the interaction between MAP/tau proteins and homologous (chick) and heterologous (Drosophila melanogaster) DNA.
  • To investigate the role of tubulin in the DNA binding properties of MAP/tau proteins.

Main Methods:

  • Isolation of MAP/tau proteins from twice-cycled chick brain microtubule preparations.
  • Nitrocellulose filter binding assays to measure DNA binding activity.
  • Equilibrium binding experiments to determine apparent affinity constants (K(app)).
  • Competition assays using homologous and heterologous DNA to assess binding specificity.

Main Results:

  • Isolated MAP/tau proteins were confirmed to be responsible for the measured DNA binding activity.
  • Two equilibrium affinity classes were identified for homologous chick DNA interaction: a high-affinity class (K(app) = 6 x 10(7) M-1) and a lower-affinity class (K(app) = 10(8) - 10(9) M-1).
  • Heterologous interaction with Drosophila melanogaster DNA showed only the lower affinity class (K(app) = 2 x 10(7) M-1).
  • Competition assays revealed the order of competitor strength as chick DNA > mouse DNA > D. melanogaster = E. coli DNA.
  • Homologous DNA was the strongest competitor, supporting previous findings.

Conclusions:

  • MAP/tau proteins exhibit conserved DNA binding interactions with specific DNA sequences.
  • These interactions are sequence-specific and do not critically depend on tubulin.
  • The findings suggest a fundamental role for MAP/tau in DNA binding, independent of microtubule polymerization.

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