Structural comparison of GLUT1 to GLUT3 reveal transport regulation mechanism in sugar porter family.

Tânia Filipa Custódio,Kelly May Frain,Peter Aasted Paulsen,Bjørn Panyella Pedersen

doi:10.26508/lsa.202000858

Abstract

The human glucose transporters GLUT1 and GLUT3 have a central role in glucose uptake as canonical members of the Sugar Porter (SP) family. GLUT1 and GLUT3 share a fully conserved substrate-binding site with identical substrate coordination, but differ significantly in transport affinity in line with their physiological function. Here, we present a 2.4 Å crystal structure of GLUT1 in an inward open conformation and compare it with GLUT3 using both structural and functional data. Our work shows that interactions between a cytosolic "SP motif" and a conserved "A motif" stabilize the outward conformational state and increases substrate apparent affinity. Furthermore, we identify a previously undescribed Cl- ion site in GLUT1 and an endofacial lipid/glucose binding site which modulate GLUT kinetics. The results provide a possible explanation for the difference between GLUT1 and GLUT3 glucose affinity, imply a general model for the kinetic regulation in GLUTs and suggest a physiological function for the defining SP sequence motif in the SP family.

Highlights

In humans, GLUT proteins are responsible for cellular glucose uptake
The results provide a provisional model which can explain the kinetic differences between GLUT1 and GLUT3, and suggests their transport regulation by a structural framework found in the Sugar Porter (SP) motif that can interact with lipids and/or intracellular sugar
The results show that expression levels are comparable (Fig S2A) and that kinetic parameters of GLUT3 are not affected by the introduced epitope (Fig S2B)

Summary

Introduction

GLUT proteins are responsible for cellular glucose uptake. MFS proteins share a common fold comprising of 12 transmembrane helices (M1–M12) with a twofold pseudo-symmetry between the N-domain (M1-6) and the C-domain (M7-12). They are defined by a signature motif, the “A motif,” with a consensus sequence of Gx3[D/E][R/K]xGx[R/K][K/R] (Nishimura et al, 1993). Due to the pseudo-symmetry, the A motif is found twice, located in the cytosolic loop connecting M2 and M3 of the N-domain and in the cytosolic loop connecting M8 and M9 of the C-domain. The SP motif takes the form of P208ESPR212 and P453ETKG457 in these two locations in the GLUT1 protein (Fig S1) (Baldwin, 1993). The importance of the SP motif has been demonstrated through mutational studies, and is highlighted by its strong conservation in the SP family (Seyfang & Landfear, 2000; Sun et al, 2012); the functional role of the SP motif has not yet been established

Methods

Results

Conclusion