User:Andy Maloney/Kinesin & Microtubule Page/Microtubule papers/Analysis of the migration behavior of single microtubules in electric fields

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Disclaimer

These are my notes. Please read the paper before reading my notes.

Paper

Analysis of the migration behaviour of single microtubules in electric fields.

Notes

  • Microtubules move towards anode (I think they meant the cathode) in an electric field with a mobility of ~ 2.6 x 10-4 cm2/V s.
    • This corresponds to an unbalanced negative charge of 0.19 electrons/tubulin dimer.
      • This is a factor of 50 difference than crystallographic data for non-assembled tubulin dimers.
    • They say that when tubulin polymerizes, it causes changes in the charge distribution of the dimer.
  • Tubulin was purified from porcine brain.
  • Kinesin was purified from porcine brain.
  • They were able to vary the field from 0 - 100 Vdc.
  • Their PEM buffer is:
    • 20 mM PIPES
    • 80 mM NaCl
    • 1 mM EGTA
    • 0.5 mM MgCl2
    • pH 6.8
    • 10 µM Taxol
    • Ionic strength = 120 mM
  • The assays used:
    • 40 µg/mL tubulin
    • 70 µg/mL kinesin
    • 5 mg/mL BSA
  • You can get the crystallographic data for tubulin for CHARMM and TINKER!
  • Microtubules disassembled when the ionic strength of the buffer was below 3 mM.
  • Microtubules have protrusions that are 12-16 amino acids long which can be as long as 4 nm.
  • Isoelectric points of tubulin:
    • alpha = 5.45 - 5.65
    • beta = 5.30 - 5.45
    • So, somewhere around these pH values, microtubules should stop moving in an electric field. But, they saw that it took a pH value of 4.1 to stop them.
  • Huh, when using Taxol, a hydrophilic cleft in beta tubulin turns into a hydrophilic surface.
  • They did not see a preferential orientation for microtubules in an electric field.
    • Wow, this is a negative result that was published.
  • They do note that when polymerizing microtubules, a pulsed electric field will align microtubules in a preferential orientation.