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Structural investigation on p-glycoprotein 1, bile salt export pump, and sodium/calcium exchanger

Küng, Raphael. Structural investigation on p-glycoprotein 1, bile salt export pump, and sodium/calcium exchanger. 2018, Doctoral Thesis, University of Basel, Faculty of Science.

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Official URL: http://edoc.unibas.ch/diss/DissB_13670

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Abstract

Transmembrane proteins span biological membranes and provide essential functions to the cell. Here we provide structural insight into three transmembrane proteins involved in substrate transport and signal transduction. Our goal is to understand the mechanism underlying the function of those proteins and to contribute thereby to improvements for future medical treatments.
P-glycoprotein 1 (Pgp) is an ABC transporter involved in multidrug transport. It provides protection from potentially toxic substances by exporting them from the cell. However in cancer cells upregulation of Pgp expression can interfere with therapy. Here we provide a near-atomic resolution structure of cross-linked nucleotide free Pgp interacting with UIC2, an inhibitory antibody, and the third generation Pgp inhibitor zosuquidar. Our structure shows binding of two zosuquidar molecules in the transmembrane domain of an occluded conformation. In addition we describe the binding interface of UIC2 binding, providing insight to the mechanism of conformational trapping. Characterized binding interfaces may be exploited for therapeutic purposes.
BSEP is the only bile salt transporter at the canalicular membrane of hepatocytes. Despite a very high sequence identity compared to Pgp, BSEP is way more selective. While diseases effected by mutations in BSEP are considerably rare, inhibition of BSEP can be the result of not intended interactions between BSEP and a variety of pharmaceutical compounds. I determined four structures of BSEP including a drug inhibited state using the antidiabetic drug glibenclamide and two structures in presence of the substrate taurocholate. These structures give rise to multiple potential models for the transport mechanism. In addition I could show that glibenclamide binds to the cytosolic facing cavity of BSEP, either repressing the connecting loop between the N- and C-terminal BSEP or locking BSEP in a inside open conformation. In addition mutations of BSEP associated with disease could be explained on structural basis.
NCX are a sodium/calcium exchangers widely spread among species, involved in calcium signalling. We have solved two crystal structures of the bacterial NCX from Thermotoga maritima in the outward facing conformation. Both conformations are substrate free, but show differences in the ion binding site. Comparison of our structures the NCX homolog from Methanococcus jannaschii suggest a simple reorientation of N-termianl helix 7 to switch NCX to the occluded state, indicating transition to the inward facing site.
Advisors:Stahlberg, Henning and Abrahams, Jan Pieter
Faculties and Departments:05 Faculty of Science > Departement Biozentrum > Structural Biology & Biophysics > Structural Biology (Stahlberg)
UniBasel Contributors:Stahlberg, Henning and Abrahams, Jan Pieter
Item Type:Thesis
Thesis Subtype:Doctoral Thesis
Thesis no:13670
Thesis status:Complete
Number of Pages:1 Online-Ressource (iv, 123 Seiten)
Language:English
Identification Number:
edoc DOI:
Last Modified:21 Aug 2020 04:30
Deposited On:20 Aug 2020 08:54

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