Selection and characterization of Affibody ligands binding to Alzheimer amyloid beta peptides

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Grönwall, CarolineKTH,Molekylär Bioteknologi (författare)

Selection and characterization of Affibody ligands binding to Alzheimer amyloid beta peptides

Artikel/kapitelEngelska2007

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Elsevier BV,2007
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LIBRIS-ID:oai:DiVA.org:kth-8127
https://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-8127URI
https://doi.org/10.1016/j.jbiotec.2006.09.013DOI

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Språk:engelska
Sammanfattning på:engelska

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Ämneskategori:ref swepub-contenttype
Ämneskategori:art swepub-publicationtype

Anmärkningar

QC 20100722
Affibody (Affibody) ligands specific for human amyloid beta (Abeta) peptides (40 or 42 amino acid residues in size), involved in the progress of Alzheimer's disease, were selected by phage display technology from a combinatorial protein library based on the 58-amino acid residue staphylococcal protein A-derived Z domain. Post-selection screening of 384 randomly picked clones, out of which 192 clones were subjected to DNA sequencing and clustering, resulted in the identification of 16 Affibody variants that were produced and affinity purified for ranking of their binding properties. The two most promising Affibody variants were shown to selectively and efficiently bind to Abeta peptides, but not to the control proteins. These two Affibody ligands were in dimeric form (to gain avidity effects) coupled to affinity resins for evaluation as affinity devices for capture of Abeta peptides from human plasma and serum. It was found that both ligands could efficiently capture Abeta that were spiked (100 microgml(-1)) to plasma and serum samples. A ligand multimerization problem that would yield suboptimal affinity resins, caused by a cysteine residue present at the binding surface of the Affibody ligands, could be circumvented by the generation of second-generation Affibody ligands (having cysteine to serine substitutions). In an epitope mapping effort, the preferred binding site of selected Affibody ligands was mapped to amino acids 30-36 of Abeta, which fortunately would indicate that the Affibody molecules should not bind the amyloid precursor protein (APP). In addition, a significant effort was made to analyze which form of Abeta (monomer, dimer or higher aggregates) that was most efficiently captured by the selected Affibody ligand. By using Western blotting and a dot blot assay in combination with size exclusion chromatography, it could be concluded that selected Affibody ligands predominantly bound a non-aggregated form of analyzed Abeta peptide, which we speculate to be dimeric Abeta. In conclusion, we have successfully selected Affibody ligands that efficiently capture Abeta peptides from human plasma and serum. The potential therapeutic use of these optimized ligands for extracorporeal capture of Abeta peptides in order to slow down or reduce amyloid plaque formation, is discussed.

Ämnesord och genrebeteckningar

TEKNIK OCH TEKNOLOGIER Industriell bioteknik hsv//swe
ENGINEERING AND TECHNOLOGY Industrial Biotechnology hsv//eng
Affibody ligand; Amyloid beta peptide; Serum depletion; Phage display; Protein engineering
Bioengineering
Bioteknik

Biuppslag (personer, institutioner, konferenser, titlar ...)

Jonsson, AndreasAffibody AB, Bromma(Swepub:kth)u1rkd9ig (författare)
Lindström, SaraKTH,Molekylär Bioteknologi(Swepub:kth)u181vco8 (författare)
Gunneriusson, ElinAffibody AB, Bromma(Swepub:kth)u1xkylew (författare)
Ståhl, StefanKTH,Molekylär Bioteknologi(Swepub:kth)u1svy8i4 (författare)
Herne, NinaAffibody AB, Bromma (författare)
KTHMolekylär Bioteknologi (creator_code:org_t)

Sammanhörande titlar

Ingår i:Journal of Biotechnology: Elsevier BV128:1, s. 162-1830168-16561873-4863

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Av författaren/redakt...: Grönwall, Caroli ...; Jonsson, Andreas; Lindström, Sara; Gunneriusson, El ...; Ståhl, Stefan; Herne, Nina

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TEKNIK OCH TEKNOLOGIER: TEKNIK OCH TEKNO ...; och Industriell biot ...

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Av lärosätet: Kungliga Tekniska Högskolan

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