Hsp33, a redox-regulated chaperone
A few years ago, our lab discovered Hsp33 as a highly efficient redox-regulated chaperone in E. coli. Hsp33 is specifically activated by reactive oxygen species (ROS) to protect cells against oxidative stress induced cell death. What makes the study of this protein so attractive is its unique functional regulation. Hsp33 is regulated on both transcriptional and posttranslational level. On transcriptional level, Hsp33 expression is regulated like any other heat shock protein, whose expression level dramatically increases once cells encounter protein-unfolding conditions (e.g., heat shock, severe oxidative stress). On posttranslational level, Hsp33’s function is regulated by reactive oxygen species (e.g., H2O2). This dual regulation makes Hsp33 inactive as a chaperone under non-stress conditions. Upon exposure of cells to reactive oxygen species that cause protein unfolding, Hsp33’s chaperone function is rapidly activated. In its activated form, Hsp33 protects the vast majority of unfolding proteins against non-specific aggregation both in vivo and in vitro.
Understanding the mechanism of Hsp33’s post-translational regulation is one of the main goals in the Jakob lab. We discovered that Hsp33 contains a redox sensor domain with four absolutely conserved cysteines, whose redox status directly controls the chaperone function of Hsp33. In vitro studies revealed that under non-stress conditions, the four cysteines are reduced and coordinate one zinc ion. This confers considerable structural stability to the sensor domain, which is compactly folded and maintains Hsp33 in an inactive conformation. Upon exposure of Hsp33 to reactive oxygen species, the cysteines form two intramolecular disulfide bonds and zinc is released (see figure, yellow domain). While this redox event is absolutely crucial for the activation of Hsp33, recent experiments have shown that it is not sufficient. We have now discovered that Hsp33 harbors an additional folding sensor region (see figure, green domain), which responds to unfolding conditions with dramatic structural rearrangements that lead to the unfolding of Hsp33’s C-terminus. Only upon the conversion of Hsp33’s C-terminus into a natively unfolded polypeptide, Hsp33 dimerizes and becomes an active molecular chaperone. This makes Hsp33 the first chaperone, whose activation mechanism involves oxidative domain unfolding in order to protect cells against oxidative protein unfolding. This strict regulation of Hsp33 reflects the in vivo requirement for Hsp33, which appears to compensate for the loss of activity of other E. coli chaperones that have been shown to become rapidly inactivated by these oxidative stress conditions.
