Biology Reference
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FIGURE 14.2
Polymerization timecourses in four different buffers. The first 1000 s of polymerization,
monitored by OD 350 , is shown for four buffers from Table 14.1 . Pipes-Mg
þ
1 M glycerol is
indicated by the dashed and dotted line, Pipes-Mg
þ
1 M TMAO by the dashed line, 1 M
glutamate by the dotted line, and Pipes-Mg
OD,
determined by subtracting the initial OD corresponding to the buffer and soluble tubulin from
all subsequent readings, is plotted versus time showing polymerization kinetics of the
systems. (B) The same data are plotted as
þ
10
m
M paclitaxel by the solid line. (A) The
D
D
OD normalized to the steady state (plateau) OD
for each buffer, which is
OD max . The OD max was taken at 30 min (1800 s) of
polymerization and is not shown on these graphs. The normalized graph shows more clearly
the differences in lag times as well as differences in the rates at which polymerization occurs
as each system approaches its maximum amount of polymer under the different buffer
conditions.
D
OD/
D
than 0.24 at 350 nm and 1 cm pathlength ( Correia & Williams, 1983 ). A solution
with sheet polymers of tubulin will have a higher OD than this (as well as characteristic
changes in a plot of OD vs. wavelength ( Detrich et al., 1985; Hall & Minton, 2005 )).
Thus simply calculating
* will allow a quick check of whether the polymerization
reaction is yielding MT or, for example, a mixture of MT and sheets. An example
of this is the polymerization of tubulin in 1 M glutamate shown in Fig. 14.2 and
e
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