(D)-p-Hydroxyphenylglycine production by thermostable D-hydantoinase from Brevibacillus parabrevis-PHG1

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.

Keywords::

• D-hydantoinase
• Brevibacillus parabrevis
• D-p-hydroxyphenylglycine
• manganese ion

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.