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Efficient protein binders for the C-reactive protein from a designed chemically modified peptide library
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
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(English)Manuscript (Other academic)
Abstract [en]

A polypeptide conjugate synthesized by coupling a small organic molecule to the side chain of an amino acid residue in a designed 42-residue polypeptide binds the C-reactive protein (CRP) essentially irreversibly. The specificity in human serum is equal to that of an avian antibody although the size is only 1/30 and the structure unordered. The polypeptide conjugate binds CRP several orders of magnitude more tightly than the small molecule due to the fact that one amino acid has been modified to include a more strongly interacting side chain. The polypeptide was selected from a 16-membered set of sequences with no prior relationship to the target protein and designed to fold into a helix-loop-helix motif. The results suggest that synthetic amino acid alphabets with more strongly interacting side chains can be used to form polypeptides with improved binding properties in comparison to those engineered by biological methods.

National Category
Natural Sciences
Identifiers
URN: urn:nbn:se:uu:diva-101403OAI: oai:DiVA.org:uu-101403DiVA: diva2:212936
Available from: 2009-04-25 Created: 2009-04-25 Last updated: 2013-07-12Bibliographically approved
In thesis
1. Polypeptide Conjugates as High-affinity Binders for Proteins
Open this publication in new window or tab >>Polypeptide Conjugates as High-affinity Binders for Proteins
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

A novel concept for protein recognition has been developed. The recognition unit is a hybrid molecule obtained by conjugation of a small organic molecule to a synthetic polypeptide selected from a 16-membered set of 42 amino acid residue sequences. The sequences are unordered and have no prior relation to the target proteins. The concept is based on the hypothesis that a small set of sequences capable of hydrophobic interactions, hydrogen bonding and electrostatic interactions can yield a binder for any selected protein, provided that the small molecule shows medium affinity or better and is reasonably selective.

The concept has been illustrated by the design, synthesis and evaluation of binders for three different proteins, the C-reactive protein, CRP, human Carbonic anhydrase II, HCAII, and Acetylcholine esterase, AChE. Highly efficient binders for CRP have been developed by conjugation of a derivative of the natural ligand, phosphocholine, to the side chain of one of the amino acids in each polypeptide. The binders in the set show a wide range of affinities for CRP and the tightest binder, 4-C10L17-PC6, binds almost irreversibly. Selected binders have been evaluated in human serum, where they capture CRP with high selectivity.High-affinity binders have been developed for HCAII, and the selectivity evaluated by extraction of the protein from blood. The binder 4-C37L34-B, a polypeptide conjugated to a spacered benzenesulphonamide residue, was able to extract Carbonic anhydrases specifically and to discriminate between the two isoforms of human Carbonic anhydrase. The conjugation of an acridine derivative to a polypeptide via a 14 atom spacer has been shown to yield a binder with high affinity and selectivity for AChE. The selectivity was demonstrated by extraction of AChE from Cerebrospinal fluid.

This thesis focuses on the development of a fast and reliable procedure for the construction, selection and evaluation of protein binders, with the ambition to develop a technology that is applicable to the development of binders for all proteins.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2009. 47 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 650
Series
Keyword
Protein recognition, polypeptide conjugates, peptide, molecular interactions, C-reactive protein, Acetylcholine esterase, Human Carboinc anhydrase
National Category
Chemical Sciences
Research subject
Organic Chemistry
Identifiers
urn:nbn:se:uu:diva-101406 (URN)978-91-554-7542-0 (ISBN)
Public defence
2009-06-12, B:21, BMC, Husargatan 3, Uppsala, 10:15 (English)
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Supervisors
Available from: 2009-05-20 Created: 2009-04-25 Last updated: 2009-05-20Bibliographically approved

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Danielson, U. Helena

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