Biomimetic self-assembly of lipase-zeolitic imidazolate frameworks with enhanced biosensing of protox inhibiting herbicides.

Protox inhibiting herbicides such as nitrofen have detrimental effects on the environment and human health. The current work aims to fabricate a Candida rugosa lipase (CRL)-based electrochemical sensor for rapid and sensitive detection of protox inhibiting herbicides (nitrofen). We proposed the use of poly(vinylpyrrolidone) (PVP) and amino-acids to promote accumulation of Zn2+ ions at the surfaces of Candida rugosa lipase (CRL) and subsequently induce self-assembly of a CRL-zeolitic imidazolate framework (ZIF) structure. This process can be easily and rapidly achieved via a one-pot facile self-assembly method. Steady-state fluorescence spectroscopy indicated that CRL has undergone a conformational change following encapsulation within the ZIF structure. This conformational change is beneficial as the prepared PVP/Glu/CRL@ZIF-8 exhibited enhanced catalytic activity (207% of native CRL), and higher substrate affinity (lower Km than native CRL) and showed high stability under harsh denaturing conditions. PVP/Glu/CRL@ZIF-8 was finally used for electrochemical biosensing of nitrofen. The fabricated biosensor has a wide linear detection range (0-100 µM), a lower limit of detection and a good recovery rate.

Enhanced catalytic stability of lipase immobilized on oxidized and disulfide-rich eggshell membrane for esters hydrolysis and transesterification.

Eggshell membrane (ESM) is an industrial waste that is available in abundance from food industry. Present study investigated the physicochemical properties of oxidized ESM and compared the efficiency of ESM and oxidized ESM as carrier for Burkholderia cepacia lipase (BCL) used in esters hydrolysis and transesterification. Following oxidation treatment, FTIR analysis and Ellman's assay showed amino acid cysteine in ESM was oxidized to form disulfide bond-containing cystine. In addition, AFM analysis showed ESM which exhibited a highly porous filamentous structure appeared to be coalesce following oxidation treatment. Oxidized ESM also showed reduced porosity (38.67%) in comparison to native ESM (51.65%). BCL were successfully immobilized on oxidized ESM through carrier activation method (enzyme loading of 5.01mg protein/g oxidized ESM). These immobilized lipase demonstrated significantly (P<0.05) enhanced catalytic stability with close to 100% of initial hydrolysis (12.03±0.29mmol/min/g) activity; and more than 85% of its initial transesterification (7.83±0.05) activity for at least 10 consecutive runs. Enhanced catalytic stability of BCL immobilized on oxidized ESM might be due to stabilization of the protein structure in oxidized ESM by disulfide bonds which helped formation of a stable bonding with BCL.