Skip Navigation


PEDS Advance Access originally published online on January 31, 2008
Protein Engineering Design and Selection 2008 21(4):247-255; doi:10.1093/protein/gzm090
This Article
Right arrow Full Text Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow All Versions of this Article:
21/4/247    most recent
gzm090v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrow Search for citing articles in:
ISI Web of Science (4)
Right arrowRequest Permissions
Google Scholar
Right arrow Articles by Kronqvist, N.
Right arrow Articles by Ståhl, S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Kronqvist, N.
Right arrow Articles by Ståhl, S.
Related Collections
Right arrow 2008
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

© The Author 2008. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org

A novel affinity protein selection system based on staphylococcal cell surface display and flow cytometry

Nina Kronqvist1,{dagger}, John Löfblom1,{dagger}, Andreas Jonsson2, Henrik Wernérus1 and Stefan Ståhl1,3

1Department of Molecular Biotechnology, School of Biotechnology, Royal Institute of Technology (KTH), AlbaNova University Center, SE-106 91 Stockholm 2Affibody AB, Box 20137, SE-161 02 Bromma, Sweden

3 To whom correspondence should be addressed. E-mail: stefans{at}biotech.kth.se

Here we describe the first reported use of a Gram-positive bacterial system for the selection of affinity proteins from large combinatorial libraries displayed on the surface of Staphylococcus carnosus. An affibody library of 3 x 109 variants, based on a 58 residue domain from staphylococcal protein A, was pre-enriched for binding to human tumor necrosis factor-alpha (TNF-alpha) using one cycle of phage display and thereafter transferred to the staphylococcal host (~106 variants). The staphylococcal-displayed library was subjected to three rounds of flow-cytometric sorting, and the selected clones were screened and ranked by on-cell analysis for binding to TNF-alpha and further characterized using biosensor analysis and circular dichroism spectroscopy. The successful sorting yielded three different high-affinity binders (ranging from 95 pM to 2.2 nM) and constitutes the first selection of a novel affinity protein using Gram-positive bacterial display. The method combines the simplicity of working with a bacterial host with the advantages of displaying recombinant proteins on robust Gram-positive bacteria as well as using powerful flow cytometry in the selection and characterization process.

Keywords: affibody/cell surface display/combinatorial protein engineering/Gram-positive bacteria/Staphylococcus carnosus

Received December 12, 2007; revised December 12, 2007; accepted December 17, 2007.

{dagger} These authors contributed equally to this work.


Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?


This article has been cited by other articles:


Home page
 Protein Eng Des SelHome page
S. A. Kenrick and P. S. Daugherty
Bacterial display enables efficient and quantitative peptide affinity maturation
Protein Eng. Des. Sel., November 10, 2009; (2009) gzp065v1.
[Abstract] [Full Text] [PDF]


Disclaimer: Please note that abstracts for content published before 1996 were created through digital scanning and may therefore not exactly replicate the text of the original print issues. All efforts have been made to ensure accuracy, but the Publisher will not be held responsible for any remaining inaccuracies. If you require any further clarification, please contact our Customer Services Department.