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Molecular recognition of proteins by functionalized folded polypeptide receptors
Linköping University, Department of Physics, Measurement Technology, Biology and Chemistry. Linköping University, The Institute of Technology.
2004 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis describes the design, synthesis and characterization of synthetic receptor molecules for the recognition and binding of proteins with applications in bioseparation and biosensing. A 42-residue polypeptide, designed to fold into a helix-loop-helix motif and dimerize in solution to form a four-helix bundle, was used as the scaffold. In the first part of the thesis it was functionalized by the incorporation of three substituents at the side chains of lysine residues. A library of 343 receptors was created and screened for affinity towards the human IgG fab fragment using SPR technology. The scaffold was reacted in a stepwise and combinatorial procedure with seven active esters in a pH controlled site-selective acylation reaction to form amides at the side chains of three lysine residues. Four receptor candidates were found to have 0.1 mM affinities and were selected for further investigation.

Both the unfunctionalized scaffold and the four selected receptors were found to bind well also to HCA II and the molecular interactions with this target protein were studied in detail. NMR studies of their interactions with 15N-labeled HCA II revealed that the peptides bound to a hydrophobic patch near the active site cleft, and SPR studies of modified receptor polypeptides led to the conclusion that mainly hydrophobic interactions were involved in binding.

In the second part of the thesis two scaffolds were functionalized with a benzenesulfonamide ligand linked to the scaffold by a series of aliphatic spacers of varying length. Benzenesulfonamide is a known inhibitor of HCA II with a dissociation constant of 1.5 µM and it was found that the overall affinity of the functionalized peptide was enhanced by increasing the length of the ligand spacer due to cooperativity between the scaffold and the ligand in the binding to HCA II. The receptor with a seven methylene group spacer bound HCA II with a dissociation constant of 4 n M. It was also shown that the sequence of the scaffold polypeptide strongly affected the overall affinity of the peptide conjugate for the target protein.

Place, publisher, year, edition, pages
Linköping: Linköpings universitet , 2004. , p. 43
Series
Linköping Studies in Science and Technology. Thesis, ISSN 0280-7971 ; 1078
National Category
Cell and Molecular Biology
Identifiers
URN: urn:nbn:se:liu:diva-153049Local ID: LiU-TEK-LIC-2004:07ISBN: 9173739170 (print)OAI: oai:DiVA.org:liu-153049DiVA, id: diva2:1278460
Available from: 2019-01-14 Created: 2019-01-14 Last updated: 2019-05-10Bibliographically approved

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Andersson, Theresa

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  • apa
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