Global S&T Development Trend Analysis Platform of Resources and Environment
Elucidating the mechanism of sugar import requires a molecular understanding of how transporters couple sugar binding and gating events. Whereas mammalian glucose transporters (GLUTs) are specialists(1), the hexose transporter from the malaria parasite Plasmodium falciparum PfHT1(2,3) has acquired the ability to transport both glucose and fructose sugars as efficiently as the dedicated glucose (GLUT3) and fructose (GLUT5) transporters. Here, to establish the molecular basis of sugar promiscuity in malaria parasites, we determined the crystal structure of PfHT1 in complex with d-glucose at a resolution of 3.6 angstrom. We found that the sugar-binding site in PfHT1 is very similar to those of the distantly related GLUT3 and GLUT5 structures(4,5). Nevertheless, engineered PfHT1 mutations made to match GLUT sugar-binding sites did not shift sugar preferences. The extracellular substrate-gating helix TM7b in PfHT1 was positioned in a fully occluded conformation, providing a unique glimpse into how sugar binding and gating are coupled. We determined that polar contacts between TM7b and TM1 (located about 15 angstrom from d-glucose) are just as critical for transport as the residues that directly coordinate d-glucose, which demonstrates a strong allosteric coupling between sugar binding and gating. We conclude that PfHT1 has achieved substrate promiscuity not by modifying its sugar-binding site, but instead by evolving substrate-gating dynamics.
Crystal structure of the Plasmodium falciparum hexose transporter PfHT1 reveals the molecular basis of its ability to transport multiple types of sugar as efficiently as the dedicated mammalian glucose and fructose transporters.
The ability to reverse protein aggregation is vital to cells(1,2). Hsp100 disaggregases such as ClpB and Hsp104 are proposed to catalyse this reaction by translocating polypeptide loops through their central pore(3,4). This model of disaggregation is appealing, as it could explain how polypeptides entangled within aggregates can be extracted and subsequently refolded with the assistance of Hsp70(4,5). However, the model is also controversial, as the necessary motor activity has not been identified(6-8) and recent findings indicate non-processive mechanisms such as entropic pulling or Brownian ratcheting(9,10). How loop formation would be accomplished is also obscure. Indeed, cryo-electron microscopy studies consistently show single polypeptide strands in the Hsp100 pore(11,12). Here, by following individual ClpB-substrate complexes in real time, we unambiguously demonstrate processive translocation of looped polypeptides. We integrate optical tweezers with fluorescent-particle tracking to show that ClpB translocates both arms of the loop simultaneously and switches to single-arm translocation when encountering obstacles. ClpB is notably powerful and rapid
A combination of optical tweezers and fluorescent-particle tracking is used to dissect the dynamics of the Hsp100 disaggregase ClpB, and show that the processive extrusion of polypeptide loops is the mechanistic basis of its activity.
Class B G-protein-coupled receptors are major targets for the treatment of chronic diseases, including diabetes and obesity(1). Structures of active receptors reveal peptide agonists engage deep within the receptor core, leading to an outward movement of extracellular loop 3 and the tops of transmembrane helices 6 and 7, an inward movement of transmembrane helix 1, reorganization of extracellular loop 2 and outward movement of the intracellular side of transmembrane helix 6, resulting in G-protein interaction and activation(2-6). Here we solved the structure of a non-peptide agonist, TT-OAD2, bound to the glucagon-like peptide-1 (GLP-1) receptor. Our structure identified an unpredicted non-peptide agonist-binding pocket in which reorganization of extracellular loop 3 and transmembrane helices 6 and 7 manifests independently of direct ligand interaction within the deep transmembrane domain pocket. TT-OAD2 exhibits biased agonism, and kinetics of G-protein activation and signalling that are distinct from peptide agonists. Within the structure, TT-OAD2 protrudes beyond the receptor core to interact with the lipid or detergent, providing an explanation for the distinct activation kinetics that may contribute to the clinical efficacy of this compound series. This work alters our understanding of the events that drive the activation of class B receptors.
