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Figure 5.
Fig. 5. GAP activity of S. pombe rna1p in the presence of
increasing concentrations of the C-terminally truncated mutant
protein rna1  p341. 1
µM Ran loaded with [  -32P]GTP
was incubated at 30 °C with buffer (  circle ),
0.025 nM rna1p (  ), 0.05 nM
rna1p (  ), or 0.1
nM rna1p (  ). In the
latter case (1^ µM Ran, 0.1 nM rna1p) the reaction was
carried out in the absence^ ( ) or
presence of 0.025 nM rna1p 341 (  ), 0.05 nM
rna1p 341 (  ), 0.1 nM
rna1p 341
(×), 0.2 nM rna1p 341 (  ), 0.5 nM
rna1p 341 (  ), 2 nM
rna1p 341 (  ), or 20 nM
rna1p 341 (  ). GTPase
activity was determined as decrease of the nitrocellulose-bound
[ -32P]GTP,
and results are given as the percentage of radioactivity bound
to Ran 30 s after addition of the respective GAP. The individual
experiments were carried out at least three times, and curves of
a representative experiment are shown. Note that the addition of
increasing amounts ot the rna1p 341 does
not affect the GAP activity of 0.1 nM wild-type rna1p. All
curves characterizing the reactions containing 1 µM Ran
and 0.1 nM rna1p are nearly identical and partially mask one
another.
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