Prolonged self-assembly of H. pylori ferritin globules at physiological conditions.

Abstract

One of the promising drug delivery tools is ferritin, which features high stability at a wide range of conditions and protects cargo by its spherical protein shell. We studied the self-assembly into homoglobules of ferritin from H. pylori and a chimeric protein ferritin-SUMO. We exposed the globules to pH-driven dis/reassembly and in both cases we observed two fractions during size exclusion chromatography (SEC) procedure. The higher molecular weight fraction contained fully assembled globules of ferritin and ferritin-SUMO that well coincides with literature. Interestingly, the lower molecular weight fraction contained intermediate subglobular oligomers that also formed globules, but on a time scale of hours, while being under physiological conditions. We performed biochemical characterization of this fraction and found that, in the case of ferritin, it contained almost the whole range of intermediate oligomers with different stoichiometry. In contrast, the ferritin-SUMO fraction contained only two distinct states: dimers and globules, without any other ferritin-SUMO intermediate oligomers. We built AlphaFold-derived schemes of ferritin and ferritin-SUMO self-assembly which also indicated differences in their assembly pathways. Our results could potentially open the possibility of cargo loading into ferritins at physiological conditions and improved purification of ferritin-based products if using a ferritin-SUMO modification with following cleavage of the SUMO-tag by the SUMO protease.