ABSTRACT
The depletion of stem cell pools and the accumulation of senescent cells in animal tissues are linked to aging. Planarians are invertebrate flatworms and are unusual in that their stem cells, called neoblasts, are constantly replacing old and dying cells. By eliminating neoblasts in worms via irradiation, the biological principles of aging are exposed in the absence of wound healing and regeneration, making planaria a powerful tool for aging research.
ABSTRACT
Ubiquitin-like modifications are important mechanisms in cellular regulation, and are carried out through several steps with reaction intermediates being thioester conjugates of ubiquitin-like proteins with E1, E2, and sometimes E3. Despite their importance, a thorough characterization of the intrinsic stability of these thioester intermediates has been lacking. In this study, we investigated the intrinsic stability by using a model compound and the Ubc9 approximately SUMO-1 thioester conjugate. The Ubc9 approximately SUMO-1 thioester intermediate has a half life of approximately 3.6 h (hydrolysis rate k = 5.33 +/- 2.8 x10(-5) s(-1)), and the stability decreased slightly under denaturing conditions (k = 12.5 +/- 1.8 x 10(-5) s(-1)), indicating a moderate effect of the three-dimensional structural context on the stability of these intermediates. Binding to active and inactive E3, (RanBP2) also has only a moderate effect on the hydrolysis rate (13.8 +/- 0.8 x 10(-5) s(-1) for active E3 versus 7.38 +/- 0.7 x 10(-5) s(-1) for inactive E3). The intrinsically high stability of these intermediates suggests that unwanted thioester intermediates may be eliminated enzymatically, such as by thioesterases, to regulate their intracellular abundance and trafficking in the control of ubiquitin-like modifications.
