Small heat shock proteins (sHsps) endow cells with stress tolerance. Of the various sHsps in mammals, HspB1, also known as Hsp27, is the most ubiquitous. To examine the structure and function of HspB1, we expressed, purified, and characterized HspB1 from Chinese hamster (Cricetulus griseus) ovary cells (CgHspB1). CgHspB1 forms a large oligomeric structure. We observed a monodisperse 16‐mer with an elongated sphere, but this is affected by changes in various conditions, including temperature. Under dilute conditions, CgHspB1 dissociates into small oligomers at elevated temperatures. The dissociated conformers interacted with the gel filtration column through hydrophobic interactions. In contrast, dissociation of the oligomer was not observed by small‐angle X‐ray scattering at 55 °C. The result partially coincides with the results of size exclusion chromatography, showing that dissociation did not occur at high protein concentrations. However, a significant structural change in the oligomeric conformations appears to occur between room and higher temperatures. Reflecting their status as homeotherms, mammalian sHsps are regulated by phosphorylation. A phosphorylation mimic mutant of CgHspB1 with the replacement of Ser15 to Asp exhibited relatively lower oligomer stability and greater protective ability against thermal aggregation than the wild‐type protein. The result clearly shows a correlation between oligomer dissociation and chaperone activity.
Plasmodium falciparum parasitophorous vacuolar protein 1 (PfPV1), a protein unique to malaria parasites, is localized in the parasitophorous vacuolar (PV) and is essential for parasite growth. Previous studies suggested that PfPV1 cooperates with the Plasmodium translocon of exported proteins (PTEX) complex to export various proteins from the PV. However, the structure and function of PfPV1 have not been determined in detail. In this study, we undertook the expression, purification, and characterization of PfPV1. The tetramer appears to be the structural unit of PfPV1. The activity of PfPV1 appears to be similar to that of molecular chaperones, and it may interact with various proteins. PfPV1 could substitute CtHsp40 in the CtHsp104, CtHsp70, and CtHsp40 protein disaggregation systems. Based on these results, we propose a model in which PfPV1 captures various PV proteins and delivers them to PTEX through a specific interaction with HSP101.
HspB1 is a mammalian sHsp that is ubiquitously expressed in almost all tissues and involved in regulating many vital functions. Although the recent crystal structure of human HspB1 showed that 24 monomers form the oligomeric complex of human HspB1 in a spherical configuration, the molecular architecture of HspB1 is still controversial. In this study, we examined the oligomeric structural change of CgHspB1 by sedimentation velocity analytical ultracentrifugation. At the low temperature of 4 °C, CgHspB1 exists as an 18-mer, probably a trimeric complex of hexamers. It is relatively unstable and partially dissociates into small oligomers, hexamers, and dodecamers. At elevated temperatures, the 24-mer was more stable than the 18-mer. The 24-mer is also in dynamic equilibrium with the dissociated oligomers in the hexameric unit. The hexamer further dissociates to dimers. The disulfide bond between conserved cysteine residues seems to be partly responsible for the stabilization of hexamers. The N-terminal domain is involved in the assembly of dimers and the interaction between hexamers. It is plausible that CgHspB1 expresses a chaperone function in the 24-mer structure.
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