Role Transformation of HSPA8 to Heme-peroxidase After Binding Hemin to Catalyze Heme Polymerization.

Hemin, a byproduct of hemoglobin degradation, inflicts oxidative insult to cells. Following its accumulation, several proteins are recruited for heme detoxification with heme oxygenase playing the key role. Chaperones play a protective role primarily by preventing protein degradation and unfolding. They also are known to have miscellaneous secondary roles during similar situations. To discover a secondary role of chaperones during heme stress we studied the role of the chaperone HSPA8 in the detoxification of hemin. In-silico studies indicated that HSPA8 has a well-defined biophoric environment to bind hemin. Through optical difference spectroscopy, we found that HSPA8 binds hemin through its N-terminal domain with a K value of 5.9 ± 0.04 µM and transforms into a hemoprotein. The hemoprotein was tested for exhibiting peroxidase activity using guaiacol as substrate. The complex formed reacts with HO and exhibits classical peroxidase activity with an ability to oxidize aromatic and halide substrates. HSPA8 is dose-dependently catalyzing heme polymerization through its N-terminal domain. The IR results reveal that the polymer formed exhibits structural similarities to β-hematin suggesting its covalent nature. The polymerization mechanism was tested through optical spectroscopy, spin-trap, and activity inhibition experiments. The results suggest that the polymerization occurs through a peroxidase-HO system involving a one-electron transfer mechanism, and the formation of free radical and radical-radical interaction. It highlights a possible role of the HSPA8-hemin complex in exhibiting cytoprotective function during pathological conditions like malaria, sickle cell disease, etc.