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A truncated and dimeric format of an Affibody library on bacteria enables FACS-mediated isolation of amyloid-beta aggregation inhibitors with subnanomolar affinity
KTH, School of Biotechnology (BIO), Protein Technology.ORCID iD: 0000-0002-5192-7362
KTH, School of Biotechnology (BIO), Protein Technology.ORCID iD: 0000-0001-9423-0541
KTH, School of Biotechnology (BIO), Protein Technology.ORCID iD: 0000-0002-9282-0174
2015 (English)In: Biotechnology Journal, ISSN 1860-6768, E-ISSN 1860-7314, Vol. 10, no 11, 1707-1718 p.Article in journal, News item (Refereed) Published
Abstract [en]

The amyloid hypothesis suggests that accumulation of amyloid β (Aβ) peptides in the brain is involved in development of Alzheimer's disease. We previously generated a small dimeric affinity protein that inhibited Aβ aggregation by sequestering the aggregation prone parts of the peptide. The affinity protein is originally based on the Affibody scaffold, but is evolved to a distinct interaction mechanism involving complex structural rearrangement in both the Aβ peptide and the affinity proteins upon binding. The aim of this study was to decrease the size of the dimeric affinity protein and significantly improve its affinity for the Aβ peptide to increase its potential as a future therapeutic agent. We combined a rational design approach with combinatorial protein engineering to generate two different affinity maturation libraries. The libraries were displayed on staphylococcal cells and high-affinity Aβ-binding molecules were isolated using flow-cytometric sorting. The best performing candidate binds Aβ with a KD value of around 300 pM, corresponding to a 50-fold improvement in affinity relative to the first-generation binder. The new dimeric Affibody molecule was shown to capture Aβ1-42 peptides from spiked E. coli lysate. Altogether, our results demonstrate successful engineering of this complex binder for increased affinity to the Aβ peptide.

Place, publisher, year, edition, pages
2015. Vol. 10, no 11, 1707-1718 p.
Keyword [en]
Affibody molecules, Affinity maturation, Amyloid beta, Bacterial display, Combinatorial protein engineering
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Research subject
Biotechnology
Identifiers
URN: urn:nbn:se:kth:diva-173855DOI: 10.1002/biot.201500131ISI: 000365129600007Scopus ID: 2-s2.0-84947041962OAI: oai:DiVA.org:kth-173855DiVA: diva2:855493
Note

Updated from E-pub to Published. QC 20151218

Available from: 2015-09-21 Created: 2015-09-21 Last updated: 2017-12-04Bibliographically approved
In thesis
1. Engineering of Affibody molecules targeting the Alzheimer’s-related amyloid β peptide
Open this publication in new window or tab >>Engineering of Affibody molecules targeting the Alzheimer’s-related amyloid β peptide
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2015. x, 107 p.
Series
TRITA-BIO-Report, ISSN 1654-2312 ; 2015:14
Keyword
Affibody molecules, Alzheimer’s disease, AD, amyloid beta, Aß, combinatorial protein engineering, staphylococcal surface display
National Category
Other Natural Sciences
Research subject
Biotechnology
Identifiers
urn:nbn:se:kth:diva-173864 (URN)978-91-7595-663-3 (ISBN)
Public defence
2015-10-09, D3, Lindstedtsvägen 5, KTH, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

QC 20150922

Available from: 2015-09-22 Created: 2015-09-21 Last updated: 2015-09-22Bibliographically approved

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Lindberg, HannaLöfblom, JohnStåhl, Stefan

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