A glutathione-based hydrogel and its site-selective interactions with water

Glutathione (gamma-glutamyl-cysteinyl-glycine; GSH) is ubiquitous biological tripeptide with multiple functions and possible therapeutic uses. The oxidized disulfide form (GSSG) self-assembles into fibrillar aggregates and gels in organic solvents, but not in solvent mixtures with high water content. Here, the disulfide bond has been replaced with a pyrenyl moiety in order to test the ability of GSH to direct noncovalent self-assembly in H2O, when combined with a hydrophobic driving force for aggregation. The resulting GSH-pyrene forms gels in 95% H2O:5% DMSO. The gamma-glutamyl group is critical for gelation, as it is with GSSG organo-gels, inasmuch as neither S-(pyrenyl)-cysteinyl-glycine nor the iodo-acetamido-pyrene precursor gels under any conditions studied. Circular dichroism and fluorescence spectroscopy indicate that the pyrene moieties cluster within the gels. Scanning and transmission electron microscopy reveal fibrous networks with individual strands of approximately 50-100 nm diameter. Saturation transfer difference (STD) NMR studies demonstrate that water interacts strongly with GSH-borne protons in both solution and gel states, but only the gels include water-pyrenyl interactions with significant residence times.     

Small molecule modulators of endogenous and co-chaperone-stimulated Hsp70 ATPase activity

The molecular chaperone and cytoprotective activities of the Hsp70 and Hsp40 chaperones represent therapeutic targets for human diseases such as cancer and those that arise from defects in protein folding; however, very few Hsp70 and no Hsp40 modulators have been described. Using an assay for ATP hydrolysis, we identified and screened small molecules with structural similarity to 15-deoxyspergualin and NSC 630668-R/1 for their effects on endogenous and Hsp40-stimulated Hsp70 ATPase activity. Several of these compounds modulated Hsp70 ATPase activity, consistent with the action of NSC 630668-R/1 observed previously (Fewell, S. W., Day, B. W., and Brodsky, J. L. (2001) J. Biol. Chem. 276, 910-914). In contrast, three compounds inhibited the ability of Hsp40 to stimulate Hsp70 ATPase activity but did not affect the endogenous activity of Hsp70. Two of these agents also compromised the Hsp70/Hsp40-mediated post-translational translocation of a secreted pre-protein in vitro. Together, these data indicate the potential for continued screening of small molecule Hsp70 effectors and that specific modulators of Hsp70-Hsp40 interaction can be obtained, potentially for future therapeutic use.