uu.seUppsala University Publications
Change search
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Corrigendum to "Histaminergic pharmacology of homo-oligomeric β3 γ-aminobutyric acid type A receptors characterized by surface plasmon resonance biosensor technology" (Biochemical Pharmacology (2012) 84 (341-351))
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Biochemistry.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Biochemistry.
Show others and affiliations
2012 (English)In: Biochemical Pharmacology, ISSN 0006-2952, E-ISSN 1356-1839, Vol. 84, no 11, 1541- p.Article in journal (Refereed) Published
Place, publisher, year, edition, pages
2012. Vol. 84, no 11, 1541- p.
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-187955DOI: 10.1016/j.bcp.2012.09.013ISI: 000311465900017OAI: oai:DiVA.org:uu-187955DiVA: diva2:576230
Available from: 2012-12-12 Created: 2012-12-12 Last updated: 2017-05-03Bibliographically 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
2. Revealing Secrets of Synaptic Protein Interactions: A Biosensor based Strategy
Open this publication in new window or tab >>Revealing Secrets of Synaptic Protein Interactions: A Biosensor based Strategy
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Protein interactions are the basis of synaptic function, and studying these interactions on a molecular level is crucial for understanding basic brain function, as well as mechanisms underlying neurological disorders. In this thesis, kinetic and mechanistic characterization of synaptic protein interactions was performed by using surface plasmon resonance biosensor technology. Fragment library screening against the reverse transcriptase of HIV was included, as it served as an outlook for future drug discovery against ligand-gated ion channels.

The protein-protein interaction studies of postsynaptic Ca2+ -binding proteins revealed caldendrin as a novel binding partner of AKAP79. Caldendrin and calmodulin bind and compete at similar binding sites but their interactions display different mechanisms and kinetics. In contrast to calmodulin, caldendrin binds to AKAP79 both in the presence and absence of Ca2+ suggesting distinct in vivo functional properties of caldendrin and calmodulin.

Homo-oligomeric β3 GABAA receptors, although not yet identified in vivo, are candidates for a histamine-gated ion channel in the brain. To aid the identification of the receptor, 51 histaminergic ligands were screened and a unique pharmacology was determined. A further requirement for identifying β3 receptors in the brain, is the availability of specific high-affinity ligands. The developed biosensor assay displayed sufficient sensitivity and throughput for screening for such ligands, as well as for being employed for fragment-based drug discovery.

AMPA receptors are excitatory ligand-gated ion channels, involved in synaptic plasticity, and modulated by auxiliary proteins. Previous results have indicated that Noelin1, a secreted glycoprotein, interacts with the AMPA receptor. By using biochemical methods, it was shown that Noelin1 interacts directly with the receptor. The kinetics of the interaction were estimated by biosensor analysis, thereby confirming the interaction and suggesting low nanomolar affinity. The results provide a basis for functional characterization of a novel AMPA receptor protein interaction.

The results demonstrate how secrets of synaptic protein interactions and function were revealed by using a molecular based approach. Improving the understanding of such interactions is valuable for basic neuroscience. At the same time, the technical advancements that were achieved to study interactions of ligand-gated ion channels by surface plasmon resonance technology, provide an important tool for discovery of novel therapeutics against these important drug targets.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2014. 73 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1133
Keyword
Surface plasmon resonance, biosensor, AMPA receptor, GABAA receptor, ligand-gated ion channel, A-kinase anchoring protein, caldendrin, calmodulin, HIV, fragment based drug discovery
National Category
Biochemistry and Molecular Biology Neurosciences
Research subject
Biochemistry; Neuroscience
Identifiers
urn:nbn:se:uu:diva-220879 (URN)978-91-554-8916-8 (ISBN)
Public defence
2014-05-16, B42, Husargatan 3, 751 23 Uppsala, 13:15 (English)
Opponent
Supervisors
Funder
EU, FP7, Seventh Framework Programme
Available from: 2014-04-25 Created: 2014-03-22 Last updated: 2014-04-29

Open Access in DiVA

No full text

Other links

Publisher's full text

Authority records BETA

Seeger, ChristianChristopeit, TonyDanielson, Helena

Search in DiVA

By author/editor
Seeger, ChristianChristopeit, TonyDanielson, Helena
By organisation
Biochemistry
In the same journal
Biochemical Pharmacology
Medical and Health Sciences

Search outside of DiVA

GoogleGoogle Scholar

doi
urn-nbn

Altmetric score

doi
urn-nbn
Total: 811 hits
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf