Beneficial Effect of Sugar Osmolytes on the Refolding of Guanidine Hydrochloride-Denatured Trehalose-6-phosphate Hydrolase fromBacillus licheniformis

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.

Biophysical Elucidation of Fibrillation Inhibition by Sugar Osmolytes in α-Lactalbumin: Multispectroscopic and Molecular Docking Approaches

“… 26 They are one of the most potent stabilizers for many proteins protecting them from denaturation or aggregation. 27 Many aromatic compounds are found to inhibit the formation of the amyloid but are questionable during in vivo studies owing to their toxic effects in clinical trials. 28 Although osmolytes are not protein-specific in general, however, in certain cases, osmolytes may destabilize or aggregate a specific protein.…”

Manufacturing Challenges and Rational Formulation Development for AAV Viral Vectors

“…2,88,111 Osmolytes Osmolytes are small organic compounds, widely used to stabilize proteins 151 against denaturation and aggregation. 8 Proteins in an aqueous solution exists in equilibrium between the folded (F) and unfolded (U) states. 152,153 Stabilization occurs by a preferential exclusion mechanism 121,154 where osmolytes shift the equilibrium towards the F-state.…”

Molecular and Biochemical Characterization of Salt-Tolerant Trehalose-6-Phosphate Hydrolases Identified by Screening and Sequencing Salt-Tolerant Clones From the Metagenomic Library of the Gastrointestinal Tract

“…According to current findings, trehalose-6-phosphate hydrolases are identified in E. coli ( Rimmele and Boos, 1994 ), Bacillus subtilis ( Sniezko et al, 1998 ), Bacillus sp. GP16 ( Karelov et al, 2003 ), Bacillus licheniformis ( Chuang et al, 2012 ; Ong et al, 2014 ; Chen et al, 2015 ; Lin et al, 2016 ), Lactobacillus acidophilus ( Duong et al, 2006 ), or Klebsiella pneumoniae ( Wu et al, 2011 ), among which Lactobacillus acidophilus is a probiotic microorganism existing in the human gastrointestinal environment. At present, the largest amount of reported trehalose-6-phosphate hydrolases found were from Bacillus .…”

“…The biochemical characterization research of trehalose-6-phosphate hydrolase mainly focused on the substrate specificity ( Rimmele and Boos, 1994 ), kinetic parameters ( Karelov et al, 2003 ), structure ( Ong et al, 2014 ), and the effect of sugar osmolytes on the refolding of trehalose-6-phosphate hydrolase ( Chen et al, 2015 ). For example, Karelov et al (2003) used T6P and p NPG as substrates to determine the K M and K cat values of trehalose-6-phosphate hydrolase at different pHs, temperatures, and NaCl conditions, respectively.…”