Figure 5.
FIG. 5. TrioN binds strongly to phospholipids only in the
presence of RhoG. His[6]-tagged bacterially purified TrioN was
applied to strips spotted with various phospholipids (Echelon)
and revealed with an anti-His or an anti-GST antibody as
detailed under "Experimental Procedures." All strips were
revealed under the same conditions. Each experiments was
repeated at least three times of which one representative is
shown. A, His[6]-TrioN at 0.5 µg/ml. B, His[6]-TrioN at 50
µg/ml. C, His[6]-TrioN/RhoG complex at 0.5 µg/ml. D,
RhoG deletion mutant lacking the C-terminal basic residues, RhoG
182, in complex with
His[6]-TrioN at 0.5 µg/ml. E, GST-RhoG at 0.5 µg/ml.
F, His[6]-TrioN/Rac complex at 0.5 µg/ml. G, Rac deletion
mutant lacking the C-terminal basic residues, Rac 184, in
complex with His[6]-TrioN at 0.5 µg/ml. H, His[6]-Rac at
0.5 µg/ml. I, bacterially expressed PLC- 1-PH
domain at 0.5 µg/ml. This strip serves as a positive
control and shows that our experimental procedure is reliable.
J, nomenclature of the different phospholipids spotted on the
nitrocellulose strip. Comparison of C and I shows that the
binding affinity of TrioN/RhoG to PtdIns(3,4)P[2], PA, or PtdIns
is similar to PLC- 1 binding to
PtdIns(4,5)P[2] ( 1 µM) (62).
The above figure is reprinted
by permission from the ASBMB:
J Biol Chem
(2004,
279,
37895-37907)
copyright 2004.
182, in complex with
His[6]-TrioN at 0.5 µg/ml. E, GST-RhoG at 0.5 µg/ml.
F, His[6]-TrioN/Rac complex at 0.5 µg/ml. G, Rac deletion
mutant lacking the C-terminal basic residues, Rac 
1-PH
domain at 0.5 µg/ml. This strip serves as a positive
control and shows that our experimental procedure is reliable.
J, nomenclature of the different phospholipids spotted on the
nitrocellulose strip. Comparison of C and I shows that the
binding affinity of TrioN/RhoG to PtdIns(3,4)P[2], PA, or PtdIns
is similar to PLC- 
1 µM) (62).