RECOVERY OF CYCLODEXTRIN GLUCANOTRANSFERASE (CGTase) USING IMMOBILIZED METAL CHELATING AFFINITY CHROMATOGRAPHY

Abstract

-Immobilized metal affinity chromatography (IMAC) was chosen as a method of purification for the recovery of CGTase from E. coli homogenate. E. coli harbouring the Bacillus sp. G1 gene expressed extracellularly secreted CGTase into ampicillin supplied LB broth. Culture was pre-purified using SnakeSkin dialysis tubing (3.5 MWCO) with an enzyme activity of 147.80 U/mL. Three strategies (A, B and C) were employed for the purification of CGTase using column adsorption chromatography with Ni 2+ -Sepharose resin. Strategy A employed an elution buffer of 50 mM EDTA, pH 7, Strategy B used 0.1 M imidazole, pH 7 and Strategy C employed 45 mM imidazole pH 7 as the elution buffer. Strategy C was found to be most suitable yielding a total CGTase recovery of 87.04% from an initial activity of 147.80 U/mL.

Molecular strategies to enhance the keratinase gene expression and its potential implications in poultry feed industry

“…The findings demonstrated that although Mg 2+ , Co 2+ , and Cu 2+ inhibited the secretion of CGTase, Fe 3+ may increase the activity of CGTase (11.5 U/mL) and biomass of the strain. The CGTase secretion was unaffected by the metal ions Mn 2+ and Ca 2+ ( Li et al, 2010 ; Sivapragasam and Abdullah 2015 ; Li et al, 2018 ). It is stopped growing the fermentation & the strain's growth on reaching 40th h of fermentation, the supernatant's CGTase production reached at optimum limit at 46.54U/mL ( Wang et al, 2018 ; Zhou et al, 2023 ).…”