Osmolyte-Like Stabilizing Effects of Low GdnHCl Concentrations on d-Glucose/d-Galactose-Binding Protein

Abstract

The ability of d-glucose/d-galactose-binding protein (GGBP) to reversibly interact with its ligands, glucose and galactose, makes this protein an attractive candidate for sensing elements of glucose biosensors. This potential is largely responsible for attracting researchers to study the conformational properties of this protein. Previously, we showed that an increase in the fluorescence intensity of the fluorescent dye 6-bromoacetyl-2-dimetylaminonaphtalene (BADAN) is linked to the holo-form of the GGBP/H152C mutant in solutions containing sub-denaturing concentrations of guanidine hydrochloride (GdnHCl). It was hypothesized that low GdnHCl concentrations might lead to compaction of the protein, thereby facilitating ligand binding. In this work, we utilize BADAN fluorescence spectroscopy, intrinsic protein UV fluorescence spectroscopy, and isothermal titration calorimetry (ITC) to show that the sub-denaturing GdnHCl concentrations possess osmolyte-like stabilizing effects on the structural dynamics, conformational stability, and functional activity of GGBP/H152C and the wild type of this protein (wtGGBP). Our data are consistent with the model where low GdnHCl concentrations promote a shift in the dynamic distribution of the protein molecules toward a conformational ensemble enriched in molecules with a tighter structure and a more closed conformation. This promotes the increase in the configurational complementarity between the protein and glucose molecules that leads to the increase in glucose affinity in both GGBP/H152C and wtGGBP.

Ameliorating amyloid aggregation through osmolytes as a probable therapeutic molecule against Alzheimer's disease and type 2 diabetes

“…42 Alexander et al found that G-HCL at low concentrations shows osmolyte-like stabilizing effects on D-glucose/D-galactosebinding proteins. 43 G-HCL was also found to show biphasic behavior during amyloid aggregation, increasing the aggregation kinetics at low G-HCL concentrations and decreasing the aggregation kinetics at high G-HCL concentrations. 44 Several studies on the inactivation and unfolding of related proteins have been done, leading to denaturation due to G-HCL.…”

Photophysical Properties of BADAN Revealed in the Study of GGBP Structural Transitions

“…In the concentration range of GdnHCl from 0 to 0.7 M, there is a slight increase in the fluorescence intensity of BADAN linked to the GGBP/W284C holoform (Figure 8D). Apparently, this is caused by the stabilizing effect of low concentrations of denaturant on the structure of the GGBP/W284C complex with glucose (Figure S5) [37].…”

“…Previously, we showed that the GGBP/H152C unfolding induced by chemical denaturants guanidine hydrochloride (GdnHCl) and urea recorded by intrinsic fluorescence of protein tryptophan residues and BADAN fluorescence differ significantly [36,37]. These results may be due to the fact that the microenvironment of BADAN and GGBP tryptophan residues reflects the conformational changes in different regions of the protein.…”