Statistical design of experiments for optimization of arsenate reductase production by Kocuria palustris (RJB-6) and immobilization parameters in polymer beads

Abstract

Detection and bioremediation of arsenic by bacterial strains are of immense importance. The enzyme arsenate reductase produced by microbes plays an important role for both. The enzyme, however, is not produced commercially. This study presents statistical optimization of operational parameters for the enhancement of arsenate reductase production by an arsenic tolerant bacterium Kocuria palustris (RJB-6). Plackett-Burman and response surface methodology (RSM) were applied to screen the significant variables and determine the interactive effect of the factors influencing enzyme production. The highest arsenate reductase activity of 31.86 U mL À1 was obtained under optimized conditions (pH 5.0; seed age 24 h; substrate concentration 150 mM) compared to that obtained under common process parameters (12.89 U mL À1 ). A K m value of 6.36 mM and V max of 6250 mmol per min per mg protein were achieved from an enzyme kinetics study. EDAX analysis confirmed cytoplasmic accumulation of arsenic by RJB-6.RSM was applied to evaluate the effect of various formulations of the selected polymeric matrices for the immobilization of the enzyme. Maximum enzyme activity was obtained with a combination of alginate, chitosan, and glutaraldehyde at 3%, 0.75% and 0.75% respectively. The immobilized enzyme retained its activity following 15 cycles of reuse and had storage stability of up to 30 days at 4 C. This ensured the potential application of the enzyme in large-scale remediation and detection of arsenic in a costeffective and environment friendly way.

A Novel Xylan-Polyvinyl Alcohol Hydrogel Bead with Laccase Entrapment for Decolorization of Reactive Black 5

“…Based on results obtained from such experimental designs, Response Surface Methodology (RSM) is widely used to predict the optimal condition via regression analysis due to its exceptional accuracy (Clark and Williges 1973). The application of RSM in optimization of hydrogel bead formation and enzyme immobilization have been reported recently for cyclodextrin glycosyltransferase (CGTase) immobilized in alginate-gelatin mixed gel beads (Rakmai et al 2015) and arsenate reductase immobilized in alginate-chitosan beads (Banerjee et al 2016).…”

Immobilization of Staphylococcus aureus Sortase A on Chitosan Particles and Its Applications in Peptide-to-Peptide Ligation and Peptide Cyclization

“…A main effect figure with a positive sign indicates that the high value of the corresponding variable is near optimum and a negative sign indicates that the low value of this variable is close to optimum [ 47 ]. The F value (20.91) of the model was found to be significant.…”

“…As reducing the shiff-base bond to a secondary amine had no significant improvement on enzyme stability and other characteristics, no reducing reagent was applied in the SrtA immobilization procedure, which also avoided potentially deleterious effects on the enzymes’ structures, the disulphide bond, and the peptide bond [ 46 ]. Addtionally, the process was optimized by response surface methodology (RSM) [ 47 ]. The immobilized SrtA was further characterized by Fourier transform infrared (FTIR) spectroscopy and scanning electron microscopy (SEM).…”

Detection and removal of emerging contaminants from water bodies: A statistical approach

“…The second order polynomial model equation's quality of fit is shown by the correlation coefficient (R 2 ) and adjusted R 2 . The threedimensional contour plots are used to illustrate the relationship between the experimental levels of each parameter and the resultant response from the model (Banerjee et al, 2016b).…”