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Beneficial effect of sugar osmolytes on the refolding of guanidine hydrochloride-denatured trehalose-6-phosphate hydrolase from Bacillus licheniformis.

Chen JH, Chi MC, Lin MG, Lin LL, Wang TF - Biomed Res Int (2015)

Bottom Line: These experimental results clearly indicated that sorbitol, sucrose, and trehalose at a concentration of 0.75 M improved the refolding yields of GdnHCl-denatured  BlTreA, probably due to the fact that these sugars favored the formation of tertiary architectures.ANS fluorescence intensity measurements revealed a reduction of exposed hydrophobic surfaces upon the treatment of denatured enzyme with sugar osmolytes.These observations suggest that sugar osmolytes possibly play a chaperone role in the refolding of chemically denatured BlTreA.

View Article: PubMed Central - PubMed

Affiliation: Department of Food Science and Technology, Chia-Nan University of Pharmacy and Science, Tainan City 71710, Taiwan.

ABSTRACT
The influence of three sugar osmolytes on the refolding of guanidine hydrochloride- (GdnHCl-) denatured trehalose-6-phosphate hydrolase of Bacillus licheniformis (BlTreA) was studied by circular dichroism (CD) spectra, fluorescence emission spectra, and the recovery of enzymatic activity. These experimental results clearly indicated that sorbitol, sucrose, and trehalose at a concentration of 0.75 M improved the refolding yields of GdnHCl-denatured  BlTreA, probably due to the fact that these sugars favored the formation of tertiary architectures. Far-UV CD measurements demonstrated the ability of sugar osmolytes to shift the secondary structure of GdnHCl-denatured enzyme towards near-native conformations. ANS fluorescence intensity measurements revealed a reduction of exposed hydrophobic surfaces upon the treatment of denatured enzyme with sugar osmolytes. These observations suggest that sugar osmolytes possibly play a chaperone role in the refolding of chemically denatured BlTreA.

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Final yield of BlTreA in the absence and presence of different osmolytes. The experimental data is derived from Figure 1.
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fig2: Final yield of BlTreA in the absence and presence of different osmolytes. The experimental data is derived from Figure 1.

Mentions: To evaluate the reactivation of denatured BlTreA, the protein samples were diluted with 50 mM Hepes-NaOH buffer (pH 8.0) to a final concentration of 5.0 μM. As compared to the self-renatured BlTreA, the reactivation rate of GdnHCl-denatured enzyme was highest by adding 0.75 M of sorbitol, sucrose, and trehalose into the renaturation system (Figure 1). However, the TreA activity gradually decreased upon the addition of 1.25 M sugar osmolytes. The final reactivation yields are shown in Figure 2. The experimental data indicate that sorbitol was more effective than any of the other two sugars for the reactivation of GdnHCl-denatured BlTreA. Also, it is worth to note that all these osmolytes affected the BlTreA reactivation in a concentration dependent manner.


Beneficial effect of sugar osmolytes on the refolding of guanidine hydrochloride-denatured trehalose-6-phosphate hydrolase from Bacillus licheniformis.

Chen JH, Chi MC, Lin MG, Lin LL, Wang TF - Biomed Res Int (2015)

Final yield of BlTreA in the absence and presence of different osmolytes. The experimental data is derived from Figure 1.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4309298&req=5

fig2: Final yield of BlTreA in the absence and presence of different osmolytes. The experimental data is derived from Figure 1.
Mentions: To evaluate the reactivation of denatured BlTreA, the protein samples were diluted with 50 mM Hepes-NaOH buffer (pH 8.0) to a final concentration of 5.0 μM. As compared to the self-renatured BlTreA, the reactivation rate of GdnHCl-denatured enzyme was highest by adding 0.75 M of sorbitol, sucrose, and trehalose into the renaturation system (Figure 1). However, the TreA activity gradually decreased upon the addition of 1.25 M sugar osmolytes. The final reactivation yields are shown in Figure 2. The experimental data indicate that sorbitol was more effective than any of the other two sugars for the reactivation of GdnHCl-denatured BlTreA. Also, it is worth to note that all these osmolytes affected the BlTreA reactivation in a concentration dependent manner.

Bottom Line: These experimental results clearly indicated that sorbitol, sucrose, and trehalose at a concentration of 0.75 M improved the refolding yields of GdnHCl-denatured  BlTreA, probably due to the fact that these sugars favored the formation of tertiary architectures.ANS fluorescence intensity measurements revealed a reduction of exposed hydrophobic surfaces upon the treatment of denatured enzyme with sugar osmolytes.These observations suggest that sugar osmolytes possibly play a chaperone role in the refolding of chemically denatured BlTreA.

View Article: PubMed Central - PubMed

Affiliation: Department of Food Science and Technology, Chia-Nan University of Pharmacy and Science, Tainan City 71710, Taiwan.

ABSTRACT
The influence of three sugar osmolytes on the refolding of guanidine hydrochloride- (GdnHCl-) denatured trehalose-6-phosphate hydrolase of Bacillus licheniformis (BlTreA) was studied by circular dichroism (CD) spectra, fluorescence emission spectra, and the recovery of enzymatic activity. These experimental results clearly indicated that sorbitol, sucrose, and trehalose at a concentration of 0.75 M improved the refolding yields of GdnHCl-denatured  BlTreA, probably due to the fact that these sugars favored the formation of tertiary architectures. Far-UV CD measurements demonstrated the ability of sugar osmolytes to shift the secondary structure of GdnHCl-denatured enzyme towards near-native conformations. ANS fluorescence intensity measurements revealed a reduction of exposed hydrophobic surfaces upon the treatment of denatured enzyme with sugar osmolytes. These observations suggest that sugar osmolytes possibly play a chaperone role in the refolding of chemically denatured BlTreA.

Show MeSH
Related in: MedlinePlus