Structural and energetic landscape of the human HOP-PrPC Axis: Multi-domain stabilization and peptide-mediated disruption by melittin.

Novel Insights into the Human Hsc70 Structure by Cross-Linking Mass Spectrometry and Molecular Modeling

Heat shock cognate protein 70 (Hsc70) is a 71 kDa molecular chaperone belonging to the Hsp70 family of heat shock proteins. These proteins act as ATP-dependent molecular machines that assist protein folding under both physiological and stress conditions such as hypoxia, heat shock, and pH fluctuations. In addition to general chaperone functions, Hsc70 performs specialized roles, including uncoating clathrin-coated vesicles, facilitating protein transport into organelles, and targeting proteins for lysosomal degradation. Members of the Hsp70 family are known to form dimers and higher oligomers, but the structural organization and functional relevance of these assemblies remain poorly understood. Earlier studies also suggested that J-domain proteins (JDPs) can promote Hsp70 dimerization. In this study, we used chemical cross-linking, high-resolution Fourier transform mass spectrometry (FTMS), 15N isotopic labeling, and advanced data analysis to investigate the structural organization of Hsc70 dimers. Cross-link-derived distance restraints enabled structural modeling of Hsc70 monomers and dimers using AlphaLink2. Our results reveal distinct ATP- and ADP-state dimer conformations that coexist in equilibrium. In the presence of the cochaperone DnaJB1, we observed a shift in the dimer–monomer equilibrium, accompanied by enhanced ATP hydrolysis and formation of intermediate species. These findings demonstrate that the Hsc70 dimer population is structurally heterogeneous and depends on nucleotide state and cochaperone interactions.

The Central Role and Therapeutic Potential of Cochaperone Networking of Chaperones in the Regulation of Biocondensates

How the regulatory networking of chaperones by cochaperones, particularly J domain proteins (JDPs; also called Hsp40 or DNAJ proteins), contributes to biocondensate formation and protein disaggregation to maintain proteostasis is discussed.

Chaperoning through time - defining DNAJA2 co-chaperone client selectivity via pulse-SILAC and BioID

Heat shock protein family D member 1 (HSPD1) modulates ribosome-related pathways and protein synthesis in lung adenocarcinoma.

Direct single-molecule visualization of Hsp90-mediated relief of an Hsp70-folding block

Heat shock protein 70 (Hsp70) and Hsp90 are essential molecular chaperones that cooperate to fold diverse client proteins, yet how their activities are coordinated to remodel clients remains unclear. To address this, we used a combination of single-molecule fluorescence resonance energy transfer and total internal reflection fluorescence microscopy to observe individual firefly luciferase proteins during sequential engagement with Escherichia coli Hsp70 (DnaK) and Hsp90 (HtpG). We show that HtpG reduces rebinding of DnaK to folding intermediates while still allowing engagement with misfolded clients, enabling productive refolding in the presence of typically inhibitory concentrations of DnaK. HtpG couples adenosine 5′-triphosphate binding and hydrolysis to promote progressive folding through localized compaction across multiple regions of the client, reducing misfolding and establishing native interdomain contacts. Kinetic simulations support a model whereby heterogeneous DnaK binding generates region-specific folding kinetics and conformational dynamics. This enables efficient subdomain folding by DnaK/HtpG and suggests that the number and position of DnaK binding sites on clients provide a mechanism by which proteins can harness chaperone promiscuity for optimal folding.

NMR assignments and secondary structure analysis of the human 5MP1 C-terminal domain