Biology Reference
In-Depth Information
FIGURE 12.4
Activated Gsa increases the intrinsic GTPase activity of tubulin and promote microtubule
depolymerization. (A) Single-turnover GTPase. Mutationally activated (GTPase-deficient) Gsa
(Q227L) was incubated with tubulin GTP (200 nM) for 30 min, and the tubulin GTPase rate
was determined. EC
50
¼
1.2 mM, V
max
¼
20 mmol of GTP hydrolyzed/min/mol of tubulin, and
n
H
¼
1.0. Dashed lines: 95% confidence interval for a hyperbolic fit. Tubulin (23 mM) was
incubated with GsaQL for 1 h at 30
C in the presence of 1 mM GTP. Microtubules were
pelleted by centrifugation, and the amount of tubulin in supernatant and pellet fractions were
quantified. GsaQL inhibited microtubule assembly with an IC
50
of 3.50.4 mM.
This research was originally published by
Dave et al. (2011)
at 30
C for 40 min in the presence or absence of different concentrations of Gs
a
QL- or
Gs
a
-derived peptide, or the control peptide Gt
a
. Tracking of microtubule plus ends is
carried out at 30
C by video-enhanced differential interference contrast microscopy
using an Olympus IX71 inverted microscope with a
100 (numerical aperture
¼
1.4)
