Figure 4.

OspG binds free UbcH5c and Ub with weak affinity.

A, B ¹H,¹⁵N-TROSY spectra of (A) free UbcH5c and Ub and (B) the UbcH5c-O˜Ub conjugate before and after addition of 1 molar equivalent of GST-OspG (grey and red, respectively). OspG shows weak interactions with free UbcH5c and Ub (A). The perturbed UbcH5c and Ub resonances map to the E3-binding interface of UbcH5c and the region surrounding the I44 hydrophobic surface of Ub as shown in the crystal structure. An enlarged region of the center (inset) shows linewidth broadening of the UbcH5c-O˜Ub spectrum after addition of OspG (B), illustrating a substantial increase in affinity attained via conjugation of UbcH5c and Ub subunits.