This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Yang, T. H.
Right arrow Articles by Rhee, J. S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Yang, T. H.
Right arrow Articles by Rhee, J. S.
Agricola
Right arrow Articles by Yang, T. H.
Right arrow Articles by Rhee, J. S.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, December 2004, p. 6968-6976, Vol. 70, No. 12
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.12.6968-6976.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Use of Pseudomonas putida EstA as an Anchoring Motif for Display of a Periplasmic Enzyme on the Surface of Escherichia coli

Taek Ho Yang,1 Jae Gu Pan,2 Yeon Soo Seo,1 and Joon Shick Rhee1*

Department of Biological Sciences, Korea Advanced Institute of Science and Technology,1 National Research Laboratory for Microbial Display, GenoFocus Inc., Daeduk BioCommunity, Daejeon, Republic of Korea2

Received 20 May 2004/ Accepted 17 July 2004

The functional expression of proteins on the surface of bacteria has proven important for numerous biotechnological applications. In this report, we investigated the N-terminal fusion display of the periplasmic enzyme ß-lactamase (Bla) on the surface of Escherichia coli by using the translocator domain of the Pseudomonas putida outer membrane esterase (EstA), which is a member of the lipolytic autotransporter enzymes. To find out the transport function of a C-terminal domain of EstA, we generated a set of Bla-EstA fusion proteins containing N-terminally truncated derivatives of the EstA C-terminal domain. The surface exposure of the Bla moiety was verified by whole-cell immunoblots, protease accessibility, and fluorescence-activated cell sorting. The investigation of growth kinetics and host cell viability showed that the presence of the EstA translocator domain in the outer membrane neither inhibits cell growth nor affects cell viability. Furthermore, the surface-exposed Bla moiety was shown to be enzymatically active. These results demonstrate for the first time that the translocator domain of a lipolytic autotransporter enzyme is an effective anchoring motif for the functional display of heterologous passenger protein on the surface of E. coli. This investigation also provides a possible topological model of the EstA translocator domain, which might serve as a basis for the construction of fusion proteins containing heterologous passenger domains.

* Corresponding author. Mailing address: Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 373-1, Guseong-dong, Yuseong-gu, Daejeon 305-701, Republic of Korea. Phone: 82-42-869-2613. Fax: 82-42-869-2610. E-mail: jsrhee1{at}webmail.kaist.ac.kr.

Applied and Environmental Microbiology, December 2004, p. 6968-6976, Vol. 70, No. 12
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.12.6968-6976.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Jose, J., Meyer, T. F. (2007). The Autodisplay Story, from Discovery to Biotechnical and Biomedical Applications. Microbiol. Mol. Biol. Rev. 71: 600-619 [Abstract] [Full Text]  
  • Wang, Z., Qi, Q., Wang, P. G. (2006). Engineering of Cyclodextrin Glucanotransferase on the Cell Surface of Saccharomyces cerevisiae for Improved Cyclodextrin Production.. Appl. Environ. Microbiol. 72: 1873-1877 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»