Elucidating the molecular mechanism of drug transport through structural studies of peptide transport proteins.

Lead Research Organisation: University of Oxford
Department Name: Biochemistry

Abstract

The design of drugs for oral administration is common practice in the pharmaceutical industry due to ease of patient administration, storage, distribution and patient compliance. Many drugs however are poorly absorbed in the gastrointestinal (GI) tract, the main route by which orally administered drugs enter the blood stream. One successful approach to overcome this obstacle has been the design of prodrugs. Prodrugs are modified drug compounds that target nutrient transporters in the GI tract. Of these, oligopeptide transporters are the most promising targets as they are highly expressed in the GI tract and are already responsible for the uptake of antibiotics. This prodrug strategy has already been successful in improving the oral bioavailability of a number of medically important antivirals. The role of this research proposal is to gain an understanding of the underlying mechanism by which these peptide transporters function. Such information will enable a more rational extension of the initial prodrug approach with benefits for both patients and the pharmaceutical industry alike.

Technical Summary

The proton oligopeptide cotransporter family, SLC15, is a family of integral membrane proteins that utilise the proton motive force to power the uptake of short chain peptides and peptidomimetics into a variety of cells. These proteins are the major routes of dietry nitrogen absorption from the small intestine and nitrogen re-absorption from the glomerular filtrate in the kidney. They are of broad pharmaceutical interest due to their ability to actively uptake a number of pharmaceutically important drugs, such as the beta-lactam antibiotics, certain antivirals and HIV protease inhibitors.
The aim of this proposal is to elucidate the molecular mechanism by which the SLC15 family of integral membrane proteins function and to gain a deeper understanding of drug transport and oral bioavailability. This research has significant clinical utility, as oral administration is currently the most successful route for drug administration. An understanding, at the molecular level, of how drugs are both recognised and transported across the gut epithelium will aid in more rational drug design and should have significant translational potential.
I plan to achieve these goals principally through the insights gained from X-ray crystallography. X-ray crystallography is particularly powerful in this regard as it allows currently unparalleled insights into molecular form and function. Functional studies will also be undertaken and directed by the insights obtained from the structural models. This will be a collaborative project with both eukaryotic and prokaryotic transporters being studied.
Using the expertise I have gained in the laboratory of Prof. So Iwata I am currently in the process of determining the first crystal structure of a peptide transport protein. The insights gained from this structure will make a significant contribution to our understanding of drug uptake in the human gastrointestinal tract. Through collaborations with medicinal chemists and physiologists I plan to extend this research to include the human members of this transport family. During my post-doctoral research I helped develop a rational platform for optimising membrane protein transporters for structural studies. This included screening for solubility, monodispersity and thermal stability in different detergents using small quantities of protein to allow rapid and cost effective identification of suitable targets. These methods have worked successfully for the bacterial peptide transport proteins and I now plan to utilise these methods to work on the mammalian SLC15 family members.

Publications

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Minhas GS (2019) Structural basis for prodrug recognition by the SLC15 family of proton-coupled peptide transporters. in Proceedings of the National Academy of Sciences of the United States of America

 
Description Responsive Mode
Amount £1,271,277 (GBP)
Funding ID BB/N006011/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 04/2016 
End 04/2020
 
Description Royal Society International Exchanges Scheme
Amount £11,100 (GBP)
Organisation The Royal Society 
Sector Charity/Non Profit
Country United Kingdom
Start 06/2012 
End 06/2014
 
Description Royal Society Research Grant
Amount £28,000 (GBP)
Organisation The Royal Society 
Sector Charity/Non Profit
Country United Kingdom
Start 08/2011 
End 08/2012
 
Description Royal Society Research Grant
Amount £13,500 (GBP)
Organisation The Royal Society 
Sector Charity/Non Profit
Country United Kingdom
Start 08/2011 
End 09/2012
 
Title Membrane Protein Crystallisation data 
Description We have collated together information on the crystallisation of all alpha helical membrane proteins in the Protein Data Bank upto 2012. 
Type Of Material Biological samples 
Year Produced 2012 
Provided To Others? Yes  
Impact We have developed a new membrane protein crystallisation screen that we are currently in the process of developing with a commercial company, Molecular Dimension Ltd. 
 
Description High resolution crystal structures of plasma membrane drug/peptide transporters. 
Organisation University of Kyoto
Department Graduate School of Medicine and Faculty of Medicine, Kyoto University
Country Japan 
Sector Academic/University 
PI Contribution We have provided highly pure and stable eukaryotic peptide transport proteins.
Collaborator Contribution Our partners have used these proteins to raise mono clonal antibodies for co-crystallisation studies.
Impact A number of monoclonals have been made and are currently in co-crystallisation trials.
Start Year 2012
 
Title MemAdvantage 
Description Development of an additive screen for membrane protein crystallisation. Currently marketed through a UK biotech company, Molecular Dimensions Ltd. 
IP Reference  
Protection Copyrighted (e.g. software)
Year Protection Granted 2012
Licensed Yes
Impact n/a
 
Title MemGold2: Membrane Protein Crystallisation Screen 
Description Unique screen for the purpose of identifying crystallisation conditions for membrane proteins. 
IP Reference  
Protection Copyrighted (e.g. software)
Year Protection Granted 2012
Licensed Yes
Impact n/a.
 
Title MemAdvantage 
Description Membrane Protein additive screen for crystal optimisation. 
Type Support Tool - For Fundamental Research
Current Stage Of Development Wide-scale adoption
Year Development Stage Completed 2012
Development Status Under active development/distribution
Impact still at early stage. 
URL http://www.moleculardimensions.com/shopdisplayproducts.asp?id=212&cat=Membrane+Protein+Additive+Kits
 
Title MemGold2 Crystallisation Screen for Membrane Proteins 
Description MemGold2 is the most up to date screen for the crystallisation of membrane proteins for structural study. The screen was developed in partnership with Molecular Dimensions Ltd. a UK biotech company, and is currently on sale worldwide through their distributors. This development was funded through my MRC CDA. 
Type Support Tool - For Fundamental Research
Current Stage Of Development Wide-scale adoption
Year Development Stage Completed 2012
Development Status Under active development/distribution
Impact Still at early stage, but the screen should accelerate research into medically important membrane protein targets. 
 
Company Name Molecular Dimension Ltd. 
Description http://www.moleculardimensions.com/ 
Impact Molecular Dimensions is a world leading supplier of screens, reagents and instrumentation for protein structure determination by X-ray crystallography. Including: modern crystallization screens and reagents, bacterial growth expression media, ready-to-use selenomethioine expression media, SWISSCI MRC 2 Well and 3 Well Crystallization Plates, 24-well crystallization plates, LaminexTM, cryocrystallography products, ESRF/EMBL sample changer kits and accessories, and a range of instrumentation for protein crystal growth and analysis.
Website http://www.moleculardimensions.com/