FIGURE 9.

Mutation of the calcium binding site reduces but does not eliminate the activity of AgG3z8. A, AgG3z8D1820/D1889A mutant protein displayed identical CD spectra in the presence or absence of calcium. B, specific binding of iodinated AgG3z8, AgG3z0, N4A mutant, and the D1820/D1889A mutant to C2C12 myotubes in 35-mm culture dishes is shown. Note that the D1820/D1889A mutant showed higher specific binding than AgG3z8N4A or AgG3z0. C, number of AChR clusters on myotube surface induced by the calcium-binding site mutant (at 0.1 μg/ml, ∼5 nm) is considerably lower than that induced by AgG3z8 at the same concentration. However, the mutant protein shows clear AChR clustering activity at a higher concentration (50 nm).