Zymograph profiling reveals a divergent evolution of sirtuin that may originate from Class III enzymes.

Sirtuins are a group of NAD-dependent deacylases that conserved in three domains of life and comprehensively involved in the regulation of gene transcription, chromosome segregation, RNA splicing, apoptosis, and aging. Previous studies in mammalian cells have revealed that sirtuins not only exist as multiple copies, but also show distinct deacylase activities in addition to deacetylation. However, the understanding of sirtuin zymographs in other organisms with respect to molecular evolution remains at an early stage. Here, we systematically analyze the sirtuin activities in representative species from archaea, bacteria, and eukaryotes, using both the high-performance liquid chromatography (HPLC) assay and a 7-amino-4-methylcoumarin (AMC)-based fluorogenic method. Global profiling suggests that the deacylase activities of sirtuins could be divided into three categories and reveals undifferentiated zymographs of Class III sirtuins, especially for those from bacteria and archaea. Nevertheless, initial differentiation of enzymatic activity was also observed for the Class III sirtuins at both paralog and ortholog levels. Further phylogenetic analyses support a divergent evolution of sirtuin that may originate from Class III sirtuins. Together, this work demonstrates a comprehensive panorama of sirtuin zymographs and provides new insights into the cellular specific regulation and molecular evolution of sirtuins.