Stabilizing Effect of Various Polyols on the Native and the Denatured States of Glucoamylase

Abstract

Different spectral probes were employed to study the stabilizing effect of various polyols, such as, ethylene glycol (EG), glycerol (GLY), glucose (GLC) and trehalose (TRE) on the native (N), the acid-denatured (AD) and the thermal-denatured (TD) states of Aspergillus niger glucoamylase (GA). Polyols induced both secondary and tertiary structural changes in the AD state of enzyme as reflected from altered circular dichroism (CD), tryptophan (Trp), and 1-anilinonaphthalene-8-sulfonic acid (ANS) fluorescence characteristics. Thermodynamic analysis of the thermal denaturation curve of native GA suggested significant increase in enzyme stability in the presence of GLC, TRE, and GLY (in decreasing order) while EG destabilized it. Furthermore, CD and fluorescence characteristics of the TD state at 71°C in the presence of polyols showed greater effectiveness of both GLC and TRE in inducing native-like secondary and tertiary structures compared to GLY and EG.

Polyols (Glycerol and Ethylene glycol) mediated amorphous aggregate inhibition and secondary structure restoration of metalloproteinase-conalbumin (ovotransferrin)

“…The addition of GL (15%) and EG (35%) cause blue shift (from 337 to 332 nm), ( Fig. 2B-C) suggesting the arrangement of Trp residue towards the more hydrophobic environment and this data is compatible with reported study [41].…”

Autolysis control and structural changes of purified ficin from Iranian fig latex with synthetic inhibitors

“…It found that two short chain alkanols (methanol and ethanol) were not effective in inducing significant α-helical structure in acid denatured ficin (ADF). Halogenols had higher influence in inducing the α-helical structure in ADF compared to alkanols [25]. …”

“…Evaluations were performed using UV-absorption, fluorescence and circular dichroism spectroscopy, as described earlier in ficin experiments [22,25,31,32] in 10 mM sodium phosphate buffers pH 7.0.…”

“…This may be related partly to its autolysis behavior, which may occur during purification [21]. Still ficin and similar pro-teases purification and identification are important works [22–25] because of their autolysis. Sometimes for get rid of the autolysis, ficin has been successfully purified with special method like pegylation [22].…”

“…Thus, ficin autolysis may act as an undesirable barrier for understanding its precise enzyme structure. Although, fortunately the initial studies were done on ficin crystallography and structural transitions in acid denatured ficin were studied in presence of halogenols and alkanols [23,25]. However, one of the reasons that structural ficin has not been precisely determined are its autolysis.…”