Ataxin-3, the
protein responsible for
spinocerebellar ataxia type-3, is a
cysteine protease that specifically cleaves poly-
ubiquitin chains and participates in the
ubiquitin proteasome pathway. The enzymatic activity resides in the N-terminal Josephin domain. An unusual feature of
ataxin-3 is its low enzymatic activity especially for mono-ubiquitinated substrates and short
ubiquitin chains. However, specific ubiquitination at
lysine 117 in the Josephin domain activates
ataxin-3 through an unknown mechanism. Here, we investigate the effects of K117 ubiquitination on the structure and enzymatic activity of the
protein. We show that covalently linked
ubiquitin rests on the Josephin domain, forming a compact globular moiety and occupying a
ubiquitin binding site previously thought to be essential for substrate recognition. In doing so, ubiquitination enhances enzymatic activity by locking the
enzyme in an activated state. Our results indicate that
ubiquitin functions both as a substrate and as an allosteric regulatory factor. We provide a novel example in which a conformational switch controls the activity of an
enzyme that mediates deubiquitination.