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Detection of Ligand Binding to Proteins through Observation of Hydration Water
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
2012 (English)In: Journal of Physical Chemistry B, ISSN 1520-6106, E-ISSN 1520-5207, Vol. 116, no 48, 13968-13974 p.Article in journal (Refereed) Published
Abstract [en]

Drug development is impeded by the need to design for each drug target a test that detects the binding of drug candidate molecules to the target protein. Therefore, a general method to detect ligand binding is highly desirable. Here, we present an observation toward developing such a method, which is based on monitoring a change in water absorption by infrared spectroscopy. Infrared spectroscopy has high sensitivity for water, and changes in its hydrogen bond pattern can be observed. We studied absorption changes of water upon the addition of phosphenolpyruvate or Mg2+ to pyruvate kinase. In each case, there is a decrease in the absorption of water in the 3000-3100 cm(-1) region on the low wavenumber side of the OH stretching vibration when a ligand binds to the protein. Our results suggest that the weaker water absorption is due to the release of protein-bound water into bulk water during ligand binding. This observation has high potential for drug development as well as for basic research because it can lead to a general method for detecting molecular association events that (i) is label-free, (ii) works with both binding partners being in aqueous solution, and (iii) is based on a universal process that takes place in all binding events.

Place, publisher, year, edition, pages
2012. Vol. 116, no 48, 13968-13974 p.
National Category
Biological Sciences
Identifiers
URN: urn:nbn:se:su:diva-84809DOI: 10.1021/jp307560rISI: 000311921700003OAI: oai:DiVA.org:su-84809DiVA: diva2:581468
Note

AuthorCount:3;

Available from: 2013-01-02 Created: 2013-01-02 Last updated: 2017-12-06Bibliographically approved
In thesis
1. Infrared spectroscopic studies: from small molecules to large
Open this publication in new window or tab >>Infrared spectroscopic studies: from small molecules to large
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Infrared light (IR) was first discovered by Friedrich Wilhelm Herschel in 1800. However, until 1940’s, molecular IR studies involved only water and small organic molecules, because of the long measurement times. Development Fourier transform infrared spectroscopy (FTIR) has minimized the time required to obtain data, making it possible to investigate bigger biological systems, e.g. proteins and nucleic acids.This thesis concentrates on the applications of different IR spectroscopic techniques to a variety of biological systems and development of new approaches to study complicated biological events.

The first paper in this work concerns using so-called caged compounds to study the aggregation of Alzheimer’s Aβ-peptide which is linked to the formation of neurotoxic fibrils in the brain. By adding caged-sulfate to the Aβ samples we were able to change the pH of the sample, while recording IR data and study fibril formation in a time-resolved manner. Then we used caged–ADP to study the production of ATP and creatine, mediated by creatine kinase (CK). Using CK as a helper enzyme we studied the effects of the phosphate binding on the secondary structure of SR Ca2+ATPse and determined the structural differences between two similar states Ca2E1ADP and Ca2E1ATP.

In the second part of the thesis we used ATR-FTIR spectroscopy and a specially designed dialysis setup, to develop a general method to detect ligand binding events by observing the IR absorbance changes in the water hydration shell around the molecules. The same method was used to determine the binding of DNA to the transcription factors of the E2F family. E2F proteins play main part in the gene regulatory networks that control cell development. However how they recognize their DNA-binding sites and the mechanism of binding is not well understood. By using ATR-FTIR, we observed the changes in the secondary structure of the proteins, as well as the distortions to the DNA upon E2F-DNA complex formation.

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2014. 59 p.
Keyword
Infrared spectroscopy, transcription factors, DNA, creatine kinase, CaATPase, water, ligand binding
National Category
Biophysics
Research subject
Biopharmaceutics
Identifiers
urn:nbn:se:su:diva-101077 (URN)978-91-7447-876-1 (ISBN)
Public defence
2014-03-28, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 4: Manuscript.

Available from: 2014-03-06 Created: 2014-02-24 Last updated: 2014-02-26Bibliographically approved

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