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Characterization of Affinity and Stability for the C-lobe of Calmodulin in Plasmodium falciparum.
Linköping University, Department of Physics, Chemistry and Biology.
Linköping University, Department of Physics, Chemistry and Biology.
Linköping University, Department of Physics, Chemistry and Biology.
Linköping University, Department of Physics, Chemistry and Biology.
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2016 (English)Independent thesis Basic level (degree of Bachelor), 10,5 credits / 16 HE creditsStudent thesis
Abstract [en]

Protein science gives important tools for designing efficient pharmaceuticals with specific target molecules. Calmodulin (CaM) is an essential protein existing in most species. In humans and the parasite Plasmodium falciparum the sequence differs in a few amino acids. Since the pandemic disease malaria is caused by P.falciparum, differences between human CaM and P.falciparum CaM is of interest for specific inhibition of CaM in P.falciparum.

 

In this project, differences in affinity and stability for the C-lobe of CaM (CaMC) in both human and P.falciparum was studied as a result of ligand binding and differences in thermal- and chemical stability. Furthermore, the secondary structure was examined by circular dichroism spectroscopy (CD). The stability differences between human CaMC and P.falciparum CaMC was examined by using CD and fluorescence spectroscopy. Fluorescence spectroscopy was also used when examining ligand binding with Trifluoperazine (TFP), 8-Anilinonaphthalene-1-sulfonic acid (ANS) and Artemisinin (ART). Both human CaMC and P.falciparum CaMC contains two tyrosines in their primary structure, which along with ANS was used as fluorophores when practising fluorescence spectroscopy. Human CaMC and P.falciparum CaMC were compared at different levels of structure and ligand docking by modelling.

 

Modelling with PyMOL and sequence alignment with protein BLAST showed that the structures of CaMC in human and CaMC in P.falciparum are similar, however, the primary structure differs at eleven positions whereof three of them are considered to be significant. Along with these differences in the primary structure there are other structural differences such as conformational openness, the area of the hydrophobic cleft and the structure of the reactive loops. The structural analysis performed by CD consolidates that CaMC have a similar secondary structure in human and P.falciparum. The results from the thermal and chemical stability analysis shows that both P.falciparum CaMC and human CaMC have stable structures. The fluorescence measurements of binding TFP to CaMC implies a higher binding affinity to human CaMC than P.falciparum CaMC. Moreover, further fluorescence measurements indicate a binding of ART to P.falciparum CaMC.

 

To gain a better understanding of both P.falciparum CaMC and human CaMC it would be of interest to investigate more specific ligands bound to the structures i.e. derivates of TFP and ART with lower Kd than the original TFP and ART; how they affect the stability, the structure and the activity of the C-lobe. This knowledge could be helpful in the development of malaria treatments. 

Place, publisher, year, edition, pages
2016. , p. 63
Keywords [en]
Plasmodium falciparum, malaria, calmodulin, C-lobe, stability, structure, binding affinity, Artemisinin, Trifluoperazin, ANS, modelling
National Category
Natural Sciences
Identifiers
URN: urn:nbn:se:liu:diva-134577ISRN: LITH-IFM-G-EX--16/3179--SEOAI: oai:DiVA.org:liu-134577DiVA, id: diva2:1075330
Subject / course
Chemical Biology
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Available from: 2017-02-20 Created: 2017-02-17 Last updated: 2017-02-20Bibliographically approved

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Fornander, ErikSmedéus, LydiaMattsson, ElinStrannermyr, MalinLassi, JohanSorjonen, LovisaBergqvist, JonathanDuong, Sun
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