Abstract
This study was aimed at the investigation of D-hydantoinase from newly isolated strains of bacteria for overproduction of D-p-hydroxyphenylglycine. It was also hoped to develop a D-hydantoinase with suitable physicochemical parameters to make a successful process for other D-amino acids. D-hydantoinase was isolated from a Gram positive bacterial strain PHG1, identified as Brevibacillus parabrevis based on 16S rRNA gene sequence analysis. The strain showed hydantoinase activity of 5.0 U/ml of broth with hydantoin as substrate, at a cell turbidity of 4.8 at 600 nm. The enzyme was purified to homogeneity with native and subunit molecular mass of ≈154 kDa and ≈43 kDa, respectively. The specific activity of the enzyme was found to be ≈750 μmole/min/mg. D,L-p-hydroxyphenylhydantoin was found to be the preferred substrate after unsubstituted hydantoin. The optimum temperature and pH for enzyme activity were 70°C and 8.5, respectively, with a half-life of 120 min at 70°C. Supplementing with 0.32 mM Mn2+ in the growth medium resulted in ≈3-fold increase in enzyme activity. Ninety-five percent conversion efficiency of D-hydantoinase for D,L-p-hydroxyphenylhydantoin (10% w/v) into N-carbamoyl-(D)-p-hydroxyphenylglycine, better pH-temperature stability and broad substrate specificity signify the usefulness of this enzyme for production of D-p-hydroxyphenylglycine and other D-amino acids of industrial importance.
Acknowledgements
We acknowledge Ms Soni Pandey, MTCC, IMTECH for 16S rRNA gene sequence analysis and thank the Director, IMTECH for providing the necessary facilities.
Declaration of interest:
The authors report no declarations of interest. The authors alone are responsible for the content and writing of the paper.