Elucidating the binding efficacy of β‐galactosidase on polyaniline–chitosan nanocomposite and polyaniline–chitosan–silver nanocomposite: activity and molecular docking insights

Abstract

BACKGROUND: Aspergillus oryzae -galactosidase ( -gal; EC 3.2.1.23) was physically adsorbed on polyaniline-chitosan nanocomposite (PANI-CS-NC) and polyaniline-chitosan-silver nanocomposite (PANI-CS-Ag-NC). Stability and kinetic studies were assessed, along with reusability and molecular docking interactions.RESULTS: According to transmission electron microscopy images, the sizes of the respective NCs were found to be in the range of 23-39 and 6-18 nm. The (effectiveness factor) values for PANI-CS-NC-and PANI-CS-Ag-NC-bound -gal were calculated to be 0.89 and 1.23, respectively, suggesting an enhancement in the activity of the enzyme in the presence of silver ions. Significant broadening was observed in pH and temperature profiles. Kinetic properties of PANI-CS-NC-and PANI-CS-Ag-NC-bound -gal showed lower K m , and two-and fourfold higher V max , respectively, compared to free enzyme. PANI-CS-Ag-NC-bound -gal was able to retain 94% of its initial activity after 10 repeated uses. Circular dichroism analysis revealed conformational changes in the secondary structure of protein upon immobilization. The genotoxic assessment revealed non-toxic behaviour of both free and enzyme-bound NC. The rate of lactose hydrolysis at 50 ∘ C exhibited by PANI-CS-Ag-NC bound -gal was 7% higher than that for the native enzyme. In-silico analysis showed efficient binding of -gal on PANI-CS-NCs due to interaction with residues that were exclusive of the active site of the enzyme. CONCLUSION: PANI-CS-Ag-NC-bound -gal preparation proved to be a robust nano-biocatalyst due to remarkable enhancement in catalytic activity along with excellent stability and reusability characteristics, which could be further utilized for efficient construction of nano-biosensors for producing lactose-free dairy products to feed lactose-intolerant patients. Lactose hydrolysis in batch processEnzymatic hydrolysis of lactose is a highly selective procedure which maintains the characteristics of a raw material in the J Chem Technol Biotechnol 2019; 94: 837-849

Safeguarding the catalytic activity and stability of polyaniline chitosan silver nanocomposite bound beta-galactosidase against product inhibitors and structurally related compound

“…This study is an extension to our previously published work [24,29] where we have demonstrated the immobilization strategy and stability enhancement of the immobilized enzyme over the soluble counterpart. Furthermore, in one of the studies, we obtained a remarkable increase in the catalytic activity of NC bound enzymes in the presence of essential ions found in milk [29].…”

“…Until now, various nanostructures, such as nanoparticles [16,17], nanofiber [18], nanotube [19], nanosheet [20] and most recently, nanoflowers [21] have been explored as support for the immobilization of industrially important enzymes [22]. Among them, polymer-based NCs have attracted considerable attention in the last few years due to their exceptional stability against chemical, electrical and mechanical exposure [23,24].…”

In silico approaches to investigate enzyme immobilization: a comprehensive systematic review

“…Studies that employed the docking methods are shown in Table S1 (ESI†). In a study, 79 Hex 8.0 software 80 was applied to dock PANI-CS-NC and PANI-CS-Ag-NC as supports and β-galactosidase. During docking, the enzyme was kept rigid, while the ligand rotated freely around the protein.…”

“…Enzyme-support binding sites were not located in the enzyme active site; therefore, no competition occurred between the substrate and the supports to bind to the enzyme active center. 79 In another study, 81 enzyme nanohybrid formation was evaluated using the MIB server 39 and docking method. In the latter procedure, blind docking between the enzyme and the support was performed.…”

Intolerance to Milk Lactose, Diagnostic Tests and Dietary Management: A Recent Update

“…A study on Kluyveromyces lactis found a therapeutic approach for lactose malabsorbers producing intolerance with subjective efficacy and no side effects (74). Efficient breakdown of lactose into glucose and galactose was observed after using exogenous lactase isolated from K. lactis and Aspergillus oryzae (75,76). The most effective method to cut down lactose in diet along with intake of necessary nutrients like calcium is to consume lactose-free dairy products (77).…”