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Powerful protein binders from designed polypeptides and small organic molecules: a general concept for protein recognition
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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2011 (English)In: Angewandte Chemie International Edition, ISSN 1433-7851, E-ISSN 1521-3773, Vol. 50, no 8, 1823-1827 p.Article in journal (Refereed) Published
Abstract [en]

High-affinity binders for the C-reactive protein (CRP), with dissociation constants in the pM to nM range and selectivities in human serum comparable to those of antibodies, were obtained by conjugation of 16 designed polypeptides to phosphocholine, a small molecule that binds CRP with a KDvalue of 5I . The polypeptides were not designed specifically to recognize CRP and bind by an adapted fit mechanism.

Place, publisher, year, edition, pages
2011. Vol. 50, no 8, 1823-1827 p.
Keyword [en]
bioorganic chemistry, C-reactive protein, phosphocholine, polypeptides, proteins
National Category
Chemical Sciences Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-147682DOI: 10.1002/anie.201005059ISI: 000287836200012PubMedID: 21328648OAI: oai:DiVA.org:uu-147682DiVA: diva2:400645
Available from: 2011-02-27 Created: 2011-02-27 Last updated: 2017-12-11Bibliographically approved
In thesis
1. Protein Interaction Studies with Low Molecular Weight Ligands: Applications for Drug Discovery, Basic Research and Diagnostic Tool Design
Open this publication in new window or tab >>Protein Interaction Studies with Low Molecular Weight Ligands: Applications for Drug Discovery, Basic Research and Diagnostic Tool Design
2013 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

In this thesis, the interactions between different proteins and small ligands were characterized by surface plasmon resonance spectroscopy (SPR) and fluorescence resonance energy transfer (FRET) based assays.   

For the C-reactive protein (CRP), a new type of artificial binder was identified which allows designing diagnostic assays superior to commonly used standard assays. Furthermore, an interaction study with the endogenous ligand phosphocholine revealed the importance of the avidity of pentameric CRP for the distinction of different types of lipid membranes. The interaction study with calcium showed how SPR based assays can be used to study ion-protein interactions despite the low atomic weight of ions.   

The transmembrane protease BACE1, an important drug target for Alzheimer’s disease, was immobilized to an SPR biosensor surface and embedded into a lipid membrane. An interaction study with a set of known BACE1 inhibitors showed that the transmembrane region has only minor effects on the interactions. Furthermore the pH-dependencies of the interactions were investigated and revealed new important conclusions for inhibitor design. Computer aided modelling showed that the protonation state of the aspartic dyad is dependent on the interacting inhibitor which offers new perspectives for in silico screenings.

The SPR assay developed for BACE1 was adapted to a more complex membrane protein, the pentameric β3 GABAA receptor. The assay allowed the pharmacological characterisation for histaminergic and GABAergic ligands and gave further evidence for cross-talk between the two signal transduction pathways. This study shows that the immobilisation method used for BACE1 and the ß3 GABAA receptor has the potential to become a standard method for handling membrane proteins.  

The identification of new drug leads from natural sources is a common strategy for drug discovery. A combination of SPR and FRET based activity assays were explored to increase the efficiency of this process. For HIV-1 protease, secreted aspartic protease (SAP) 1, 2 and 3 extracts from a marine vertebrate were identified containing potent inhibitors which interacted with the active site of the enzymes.

The studies in this thesis show that the investigation of protein interactions is crucial for understanding protein functions and can help to develop novel drugs for the treatment of different diseases.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2013. 34 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1007
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-188328 (URN)978-91-554-8566-5 (ISBN)
Public defence
2013-02-14, B21, BMC, Husargatan 3, Uppsala, 13:15 (English)
Opponent
Supervisors
Available from: 2013-01-24 Created: 2012-12-14 Last updated: 2013-04-02Bibliographically approved

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Christopeit, TonyDanielson, U. Helena

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