Applied Microbiology and Biotechnology

, Volume 103, Issue 21–22, pp 8839–8851 | Cite as

Biocatalytic production of D-p-hydroxyphenylglycine by optimizing protein expression and cell wall engineering in Escherichia coli

  • Yang Liu
  • Lingfeng Zhu
  • Wenpeng Qi
  • Bo YuEmail author
Biotechnological products and process engineering


D-p-hydroxyphenylglycine (D-HPG) functions as an intermediate and has important value in antibiotic industries. The high pollution and costs from chemical processes make biotechnological route for D-HPG highly desirable. Here, a whole-cell transformation process by D-hydantoinase(Hase) and D-carbamoylase(Case) was developed to produce D-HPG from DL-hydroxyphenylhydantoin(DL-HPH) in Escherichia coli. The artificially designed ribosome binding site with strong intensity significantly facilitated the protein expression of limiting step enzyme Case. Next, the cell wall permeability was improved by disturbing the peptidoglycan structure by overproduction of D,D-carboxypeptidases without obviously affecting cell growth, to increase the bioavailability of low soluble hydantoin substrate. By fine-tuning regulation of expression level of D,D-carboxypeptidase DacB, the final production yield of D-HPG increased to 100% with 140 mM DL-HPH substrate under the optimized transformation conditions. This is the first example to enhance bio-productivity of chemicals by cell wall engineering and creates a new vision on biotransformation of sparingly soluble substrates. Additionally, the newly demonstrated ‘hydroxyl occupancy’ phenomenon when Case reacts with hydroxyl substrates provides a referential information for the enzyme engineering in future.


D-p-hydroxyphenylglycine Cell permeability D,D-carboxypeptidase D-carbamoylase Escherichia coli 


Funding information

The work was supported by Beijing Natural Science Foundation, China (5182021), National Science Foundation of China (11604359), and Ministry of Science & Technology, China (KY201701011).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interests.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.CAS Key Laboratory of Microbial Physiological and Metabolic Engineering, State Key Laboratory of Mycology, Institute of MicrobiologyChinese Academy of SciencesBeijingChina
  2. 2.University of Chinese Academy of SciencesBeijingChina
  3. 3.IBG-1: Biotechnology, Institute of Bio- and GeosciencesForschungszentrum JülichJülichGermany
  4. 4.Shanghai Institute of Applied PhysicsChinese Academy of SciencesShanghaiChina
  5. 5.China-Thailand Joint Laboratory on Microbial BiotechnologyBeijingChina

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