Figure 7.

OsBBI1 is a RING-H2 protein with E3 ligase activity in vitro. (A) Amino acid alignments of the OsBBI1 RING domain with RING domains of other proteins from Arabidopsis (At), rice (Os) and maize (Zm). Filled triangles indicate the conserved cysteine (C) residues, while asterisks indicate conserved histidine (H) residues. (B, C) E3 ligase activity of OsBBI1 in vitro. His-tagged OsBBI1 or OsBBI1^ΔRING fusion protein was assayed for E3 ubiquitin ligase activity in the presence of E1, E2, and Ubi. Ubiquitinated proteins were detected by protein blot analysis using an antibody to Ubi, with time-course increase in ubiquitinated proteins shown in B. Arrow indicates the ubiquitinated proteins.