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Figure 3.
(a) Surface (CBP80) and ribbon (CBP20) representations of the
CBC (PDB 1N52 (ref. 27)) illustrate the stabilization of the
N-terminal hinge of CBP20 by residues in its binding groove
within CBP80. The N-terminal region of CBP20 (which includes
residues Ser11, Asp12 and Ser13, shown in dark blue) enters a
groove formed by residues Lys327 and Glu328 of CBP80 (green),
making contacts that stabilize residues of CBP20 in loop  2-
3
(magenta) in the cap-bound conformation. The inset shows these
interactions in detail. Residues 26–36 of the N-terminal
region of CBP80 are in black. (b) HeLa cells were transiently
transfected with constructs encoding V5-CBP80 or V5-CBP80-K327A
E328A and HA-CBP20, as indicated, for 24 h. The expressed CBP80
proteins were immunoprecipitated with anti-V5 and western
blotted with anti-HA to show that CBP20 co-imunoprecipitated
equally well with wild-type and mutated CBP80 (as quantified by
densitometry). Immunoprecipitated CBC was assayed for UV
cross-linking incorporation of [ ^32P]GTP
(c) and ^32P-labeled capped mRNA (d), with and without an excess
of cap.
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