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Figure 3.
Fig. 3. The GRK2-binding surface of G  [q]. (A)
Stereoview of the interface. The switch II and 3
helices from G [i/q] are shown
as C traces; the 5
and 6 helices from
GRK2 are shown as cartoon ribbons. Side chains of interfacial
residues are shown as ball-and-stick models, with carbon atoms
from G [i/q] and GRK2
colored cyan and yellow, respectively. Hydrogen bonds are shown
as dashed black lines. Residues targeted by site-directed
mutagenesis in this study are underlined. (B) Sequence alignment
of the switch regions and the 3/ß5
sequence for representative members of all four G subfamilies.
Switch regions (I to III) are outlined in black and are assigned
on the basis of comparison of the active and deactivated
structures of G [i1]. Secondary
structure is represented by cylinders and arrows for helices and
ß strands, respectively. G residues that
contact effectors are green, those that bind GAPs are red, and
those that contact both are purple. Contacting residues that
were chimeric (i.e., nonnative) in the crystal structures of the
G [t] and G [13] effector
complexes are shown in a lighter shade of the appropriate color.
Green boxes outline G [i] residues
proposed to interact with adenylyl cyclase (50), and asterisks
indicate conserved residues that contribute to the hydrophobic
effector-binding pocket. The crystal structures used for these
assignments are those of G [i/q]-GRK2-Gß
 (this study), G
[i]-RGS4 [Protein
Data Bank (PDB) code 1AGR [PDB]
] (12), G [t]-PDE -RGS9 (1FQJ) (8),
G [13]-p115RhoGEF
(1SHZ) (10), and G [s]-adenylyl
cyclase (1AZS) (7). The sequences are those of mouse G [q] (M55412 [GenBank]
), mouse G [11] (NP_034431
[GenBank]
), mouse G [14] (NP_032163
[GenBank]
), human G [16] (M63904 [GenBank]
), rat G [i1] (M17527 [GenBank]
), bovine G [t] (P04695 [GenBank]
), mouse G [13] (NP_034433
[GenBank]
), and bovine G [s] (M13006 [GenBank]
). (C) Mutational analysis of G [q] residues that
directly interact with GRK2. Lysates of HEK293 cells expressing
G [q] mutants were
subjected to limited trypsin digestion in the presence and
absence (shown only for wild type) of  and
immunoblotted with G [q]-specific
antibody (upper left) (31).
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