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Applied Microbiology and Biotechnology

, Volume 103, Issue 2, pp 793–806 | Cite as

Improving extracellular protein production in Escherichia coli by overexpressing D,D-carboxypeptidase to perturb peptidoglycan network synthesis and structure

  • Haiquan YangEmail author
  • Jinyuan Hu
  • Xiao Lu
  • Fuxiang Wang
  • Wei Shen
  • Wei Hu
  • Lingling Wang
  • Xianzhong Chen
  • Long Liu
Biotechnologically relevant enzymes and proteins

Abstract

Most recombinant proteins in Escherichia coli are not efficiently secreted to the extracellular space. Structural stabilisation of the cell wall is essential for extracellular protein production in E. coli, for which D,D-carboxypeptidases are essential. Herein, we perturbed the peptidoglycan structure of the E. coli cell wall by overexpressing D,D-carboxypeptidase genes dacA or dacB, and investigated the effect on extracellular protein production. Overexpression of dacA or dacB promoted the accumulation of intracellular soluble peptidoglycan, altered cell morphology (shape and size) and led to the formation of transparent globular structures in E. coli cells. Compared with controls (CK), extracellular production of recombinant green fluorescent protein (GFP) was increased by 1.7- and 2.3-fold upon overexpression of dacA and dacB, respectively. Similarly, extracellular production of recombinant amylase and α-galactosidase was increased by 4.5- and 2.8-fold, respectively, upon overexpression of dacA, and by 11.9- and 2.5-fold, respectively, upon overexpression of dacB. Overexpression of dacA or dacB enhanced both the outer and inner membrane permeability of E. coli. This cell wall engineering strategy opens up a new direction for enhancing extracellular protein and chemical production in E. coli.

Keywords

Extracellular protein production Overexpression D,D-carboxypeptidase Peptidoglycan structure Membrane permeability Escherichia coli 

Notes

Author contributions

H.Y. designed the research; H.Y., J.H., X.L., L.W., and F.W. performed the research; X.L., W.S., F.W., and X.C. analysed the data; L.L., W.H., and H.Y. wrote the paper.

Funding information

This work was funded by National Natural Science Foundation of China (21406089), Natural Science Foundation of Jiangsu Province (BK20140152), the Open Project Program of the Key Laboratory of Industrial Biotechnology, Ministry of Education, China (KLIB-KF201509), the Open Project Program of the Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, China (KLCCB-KF201607, KLCCB-KF201802), 111 Project (111-2-06), and Postgraduate Education Research and Practice Project of Jiangnan University (YJSJG2017004).

Compliance with ethical standards

This article is in compliance with ethical standards, and does not contain any studies with animals performed or human participants.

Conflict of interest

The authors declare that they have no competing interest.

Supplementary material

253_2018_9510_MOESM1_ESM.pdf (625 kb)
ESM 1 (PDF 625 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of EducationJiangnan UniversityWuxiChina
  2. 2.Institute of Modern PhysicsChinese Academy of SciencesLanzhouChina

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