Imaging microtubules and kinesin decorated microtubules using tapping mode atomic force microscopy in fluids.
The atomic force microscope has been used to investigate microtubules and kinesin decorated microtubules in aqueous solution adsorbed onto a solid substrate. The netto negatively charged microtubules did not adsorb to negatively charged solid surfaces but to glass covalently coated with the highly positively charged silane trimethoxysilylpropyldiethylenetriamine (DETA) or a lipid bilayer of 1,2-dipalmitoyl-3-dimethylammoniumpropane. Using electron beam deposited tips for microtubules adsorbed on DETA, single protofilaments could be observed showing that the resolution is up to 5 nm. Under conditions where the silane coated surfaces are hydrophobic, microtubules opened, presumably at the seam, whose stability is lower than that of the bonds between the other protofilaments. This led to a "sheet" with a width of about 100 nm firmly attached to the surface. Microtubules decorated with a stoichiometric low amount of kinesin molecules in the presence of the non-hydrolyzable ATP-analog 5'-adenylylimidodiphosphate could also be adsorbed onto silane-coated glass. Imaging was very stable and the molecules did not show any scan-induced deformation even after hundreds of scans with a scan frequency of 100 Hz.
Kacher
C M
CM
Physik Department der TU-München, Institut für Biophysik E22, James-Franck-Strasse, D-85747 Garching, Germany.
Weiss
I M
IM
Stewart
R J
RJ
Schmidt
C F
CF
Hansma
P K
PK
Radmacher
M
M
Fritz
M
M
D000818:Animals; D001921:Brain / Q000648:ultrastructure; D007202:Indicators and Reagents; D016547:Kinesins / Q000648:ultrastructure*; D008051:Lipid Bilayers; D018625:Microscopy, Atomic Force / Q000379:methods; D008870:Microtubules / Q000648:ultrastructure*; D012996:Solutions; D013552:Swine; D014867:Water
D007202:Indicators and Reagents; D008051:Lipid Bilayers; D012996:Solutions; D014867:Water; D016547:Kinesins