Scientia Iranica

Scientia Iranica

An Efficient Procedure for the Production of trans-4-Hydroxy-L-proline Using Recombinantly Expressed Proline Hydroxylase

Authors
Jiaojiao Chen 1
Dandan Gu 1
Tianyun Li 1
Jiansong Ju 1
Zhangwei Xue 2
Cunhui Li 2
Jin Yan 3
Jinxiu Zhang 1
Li-an Wang 1
1 College of Life Sciences, Hebei Normal University, No. 20, Road East of 2nd Ring South, Yuhua District, Shijiazhuang, 050024, China
2 Hebei Brant Pharmaceutical Co., LTD, Mayu Industrial Park, Jinzhou, 052260, China
3 Beijing Hongyisifang Radiation Technology Co., LTD, Beijing, 101113, China
Abstract
Due to the codon usage and high G+C contentof thetrans-4-proline-L-hydroxylase gene from the Dactylosporangium sp.strain RH1, the whole gene was optimized and cloned into several vectors for expression. In biotransformations with resting cells, the activity of the enzyme was investigated. The in-house modified plasmid pet-M-3C was found to yield the highest enzymatic activity. Additionally, after the primary fragment screening, the conversion efficiency of fragment 1-257 aa was enhanced from 76.60% to 88.97% compared with the full-length proline 4-hydroxylase within 60 h; we also found that truncation of the gene improved the solubility of the encoded protein.After optimizing the various induction conditions with respect to the enzymatic activity of the engineered strain, including the concentration of the inducer IPTG, the cell density before the inducer was added, the induction temperature and the induction time, the conversion efficiency was more than 97%within 48 h.
Keywords
proline
hydroxy-L-proline
proline hydroxylases
optimization
conversion efficiency
truncation
Scientia Iranica
Volume 22, Issue 6 - Serial Number 6
Transactions on Chemistry and Chemical Engineering (C)
December 2015
Pages 2350-2357

  • Receive Date 07 October 2015
  • Accept Date 27 July 2017