FIG. 8.

AAV2 lysine mutant K532R demonstrates reduced ubiquitination compared with the AAV2-WT and AAV5-WT vectors. (A) Approximately 3×10⁸ viral particles of AAV2-WT, AAV5-WT, and AAV2 K532R vectors were denatured at 95°C for 5 min. The denatured viral particles were then used to perform the ubiquitin conjugation assay according to the manufacturer's protocol. The processed samples were electrophoresed on a 5–20% denaturing polyacrylamide gel and the ubiquitination pattern was detected by immunoblotting with an anti-ubiquitin antibody. The mono- to-polyubiquitin conjugates were detected as a smear at molecular mass >150 kDa. (B) Capsid VP1, VP2, and VP3 proteins were used as loading control.