The overarching aim in this proposal is to determine at atomic resolution the structural basis of glucose transporter (GLUT) function. This will reveal, at a molecular level, its transport mechanism and involvement in pathologies of diabetes, cancer and other diseases. It will also provide a rational platform for developing both novel drugs and medical detection methods. GLUT carries carbohydrates (glucose, fructose, galactose and others), major energy sources for living cells, across membranes. So far fourteen human GLUTs have been identified and characterized. They exhibit significant variability in function and tissue expression despite their relatively high sequence similarity. Determining the three-dimensional (3D) structure of any GLUT will be a remarkable breakthrough in the study of this fundamental transport system that wil open new research avenues. Structural information in conjunction with site-directed mutagenesis and functional studies will allow us to pinpoint the molecular base of the functional variability of GLUT members, so that designed ligands for a particular GLUT can be rationally pursued. To understand the functional diversity of glucose transporters and increase the chances of obtaining a structural solution, the targets include: human glucose transporters (hGLUTs), and several bacterial counterparts with greater than 25% identity and 45% homology to hGLUTs amino-acid sequences. Milligram quantities of purified, functional protein for seven bacterial and three human GLUTs were produced. Thus far, crystallization efforts have been successful with four bacterial GLUTs among which two produced diffracting crystals;crystals of one bacterial GLUT diffracted up to 3.1 Angstroms resolution. Our hypotheses are that 1) the 3D structure of bacterial GLUTs will be representative for those of human GLUTs, as the active site residues are conserved from prokaryotic to eukaryotic species, and 2) as a MFS member, GLUT has two symmetric 6-helicies bundles and a hydrophilic internal cavity which undergo conformational changes during the transport mechanism. Our two specific aims are to: 1a) Determine the 3D structures of those bacterial GLUTs that yield well-diffracting crystals. 1b) Produce diffracting crystals of at least one purified human GLUT. 2a) Investigate the transport of purified GLUTs. 2b) Develop high-throughput binding assay for GLUTs in order to find novel inhibitors. The results of our proposed studies will impact our fundamental understanding of GLUTs. Furthermore, they will empower the search for novel drugs for the treatment of diseases involving GLUTs in two ways: 1) structure determination of GLUTs will guide rational drug design;2) identification of novel ligands through high- throughput screening of small molecules will provide lead compounds for new drugs.

Public Health Relevance

Glucose transporters are ubiquitous membrane proteins that carry carbohydrates (glucose, fructose, galactose and others) across cell membranes. In humans they have been implicated in pathologies of diabetes, cancer and neurological disorders. The results of the proposed studies will impact the fundamental understanding of the transporters and facilitate rational drug discovery for the treatment of various diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
1R01DK091754-01A1
Application #
8194680
Study Section
Biochemistry and Biophysics of Membranes Study Section (BBM)
Program Officer
Sechi, Salvatore
Project Start
2011-07-01
Project End
2016-05-31
Budget Start
2011-07-01
Budget End
2012-05-31
Support Year
1
Fiscal Year
2011
Total Cost
$386,250
Indirect Cost
Name
Rosalind Franklin University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
069501252
City
North Chicago
State
IL
Country
United States
Zip Code
60064
Ferraris, Ronaldo P; Choe, Jun-Yong; Patel, Chirag R (2018) Intestinal Absorption of Fructose. Annu Rev Nutr 38:41-67
Schmidl, Sina; Iancu, Cristina V; Choe, Jun-Yong et al. (2018) Ligand Screening Systems for Human Glucose Transporters as Tools in Drug Discovery. Front Chem 6:183
Tripp, Joanna; Essl, Christine; Iancu, Cristina V et al. (2017) Establishing a yeast-based screening system for discovery of human GLUT5 inhibitors and activators. Sci Rep 7:6197
Jordan, Paulina; Choe, Jun-Yong; Boles, Eckhard et al. (2016) Hxt13, Hxt15, Hxt16 and Hxt17 from Saccharomyces cerevisiae represent a novel type of polyol transporters. Sci Rep 6:23502
George Thompson, Alayna M; Ursu, Oleg; Babkin, Petr et al. (2016) Discovery of a specific inhibitor of human GLUT5 by virtual screening and in vitro transport evaluation. Sci Rep 6:24240
George Thompson, Alayna M; Iancu, Cristina V; Nguyen, Thi Thanh Hanh et al. (2015) Inhibition of human GLUT1 and GLUT5 by plant carbohydrate products; insights into transport specificity. Sci Rep 5:12804
Choe, Jun-Yong (2015) Artist's Statement: Crystallography as Art. Acad Med 90:1631
Babkin, Petr; George Thompson, Alayna M; Iancu, Cristina V et al. (2015) Antipsychotics inhibit glucose transport: Determination of olanzapine binding site in Staphylococcus epidermidis glucose/H(+) symporter. FEBS Open Bio 5:335-40
Iancu, Cristina V; Zamoon, Jamillah; Woo, Sang Bum et al. (2013) Crystal structure of a glucose/H+ symporter and its mechanism of action. Proc Natl Acad Sci U S A 110:17862-7