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Determinants of sugar-induced influx in the mammalian fructose transporter GLUT5
Stockholm University, Science for Life Laboratory (SciLifeLab). Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.ORCID iD: 0000-0002-6855-9295
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics. Stockholm University, Science for Life Laboratory (SciLifeLab).
Stockholm University, Faculty of Science, Department of Biochemistry and Biophysics.
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Number of Authors: 82023 (English)In: eLIFE, E-ISSN 2050-084X, Vol. 12, article id e84808Article in journal (Refereed) Published
Abstract [en]

In mammals, glucose transporters (GLUT) control organism-wide blood-glucose homeostasis. In human, this is accomplished by 14 different GLUT isoforms, that transport glucose and other monosaccharides with varying substrate preferences and kinetics. Nevertheless, there is little difference between the sugar-coordinating residues in the GLUT proteins and even the malarial Plasmodium falciparum transporter PfHT1, which is uniquely able to transport a wide range of different sugars. PfHT1 was captured in an intermediate 'occluded' state, revealing how the extracellular gating helix TM7b has moved to break and occlude the sugar-binding site. Sequence difference and kinetics indicated that the TM7b gating helix dynamics and interactions likely evolved to enable substrate promiscuity in PfHT1, rather than the sugar-binding site itself. It was unclear, however, if the TM7b structural transitions observed in PfHT1 would be similar in the other GLUT proteins. Here, using enhanced sampling molecular dynamics simulations, we show that the fructose transporter GLUT5 spontaneously transitions through an occluded state that closely resembles PfHT1. The coordination of D-fructose lowers the energetic barriers between the outward- and inward-facing states, and the observed binding mode for D-fructose is consistent with biochemical analysis. Rather than a substrate-binding site that achieves strict specificity by having a high affinity for the substrate, we conclude GLUT proteins have allosterically coupled sugar binding with an extracellular gate that forms the high-affinity transition-state instead. This substrate-coupling pathway presumably enables the catalysis of fast sugar flux at physiological relevant blood-glucose concentrations.

Place, publisher, year, edition, pages
2023. Vol. 12, article id e84808
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URN: urn:nbn:se:su:diva-220841DOI: 10.7554/eLife.84808ISI: 001024510300001PubMedID: 37405832Scopus ID: 2-s2.0-85163948061OAI: oai:DiVA.org:su-220841DiVA, id: diva2:1797266
Available from: 2023-09-14 Created: 2023-09-14 Last updated: 2023-11-06Bibliographically approved
In thesis
1. The molecular basis for substrate recognition and gating in sugar transporters
Open this publication in new window or tab >>The molecular basis for substrate recognition and gating in sugar transporters
2023 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Sugar is a vital sustenance for most forms of life. For a cell to take up sugar, specialized transport proteins embedded into the membrane bilayer known as sugar porters, are required. Dysfunction of sugar porters is associated with some metabolic diseases, and their expression is upregulated in many cancers as they typically require more sugar than normal cells. Furthermore, sugar porters also play a role in the vitality of the malaria parasite.

The mechanism of sugar transport is known as a rocker-switch alternating access mechanism. Simplistically, sugar binds between two similar domains on the outside of a sugar transporter and the domains then move around the sugar, so the sugar is exposed to the inside. During this domain movement, protein mass will block the sugar binding site from both outside and inside, forming the occluded state which is essential to ensure no substrate leakage during transport. Despite this relatively simple model of transport, little is known about how different sugar porters display diverse substrate specificity, affinity, and turnover.

In the four papers making up this thesis, we structurally characterize missing pieces of the sugar transport cycle, identify how these states are connected with simulations, and assess factors contributing to sugar transport by functional assays. With simulations, we show how sugar catalyzes conformational change by interacting with the occluded state. We demonstrate our functional proteoliposome-based transport assay, which allows us to measure the effect of protein mutations, inhibitors, and lipid influences in sugar recognition and turnover. Characterization of the malaria parasite hexose transporter PfHT1 has allowed us to understand antimalarial inhibitor specificity against this protein which could have implications in combating the disease, as well as pharmacological control of sugar porters in general.

Place, publisher, year, edition, pages
Stockholm: Department of Biochemistry and Biophysics, Stockholm University, 2023. p. 63
Keywords
membrane transport, sugar transporter, simulations, lipids, antimalarial drugs
National Category
Biochemistry and Molecular Biology Biophysics
Research subject
Biochemistry
Identifiers
urn:nbn:se:su:diva-222119 (URN)978-91-8014-528-2 (ISBN)978-91-8014-529-9 (ISBN)
Public defence
2023-12-08, Vivi Täckholmssalen (Q211), NPQ-huset, Svante Arrhenius väg 20 and online via Zoom, public link is available at the department website, Stockholm, 14:30 (English)
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Available from: 2023-11-15 Created: 2023-10-09 Last updated: 2023-11-07Bibliographically approved

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McComas, SarahReichenbach, TomAlleva, ClaudiaBonaccorsi, MartaDrew, David

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