Investigating the Mechanism of Substrate Uptake and Release in the Glutamate Transporter Homologue Glt_Ph through Metadynamics Simulations

A homeostatic concentration of glutamate in the synaptic cleft ensures a correct signal transduction along the neuronal network. An unbalance in this concentration can lead to neuronal death and to severe neurodegenerative diseases such as Alzheimer’s or Parkinson’s. Glutamate transporters play a crucial role in this respect because they are responsible for the reuptake of the neurotransmitter from the synaptic cleft, thus controlling the glutamate concentration. Understanding the molecular mechanism of this transporter can provide the possibility of an exogenous control. Structural studies have shown that this transporter can assume at least three conformations, thus suggesting a pronounced dynamical behavior. However, some intermediate states that lead to the substrate internalization have not been characterized and many aspects of the transporter mechanism still remain unclear. Here, using metadynamics simulations, we investigate the substrate uptake from the synaptic cleft and its release in the intracellular medium. In addition, we focus on the role of ions and substrate during these processes and on the stability of the different conformations assumed by the transporter. The present dynamical results can complement available X-ray data and provide a thorough description of the entire process of substrate uptake, internalization, and release

Discovery of Covalent Inhibitors of Glyceraldehyde-3-phosphate Dehydrogenase, A Target for the Treatment of Malaria

We developed a new class of covalent inhibitors of Plasmodium falciparum glyceraldehyde-3-phosphate dehydrogenase, a validated target for the treatment of malaria, by screening a small library of 3-bromo-isoxazoline derivatives that inactivate the enzyme through a covalent, selective bond to the catalytic cysteine, as demonstrated by mass spectrometry. Substituents on the isoxazolinic ring modulated the potency up to 20-fold, predominantly due to an electrostatic effect, as assessed by computational analysis

Development of Radiolabeled Ligands Targeting the Glutamate Binding Site of the <i>N</i>‑Methyl‑d‑aspartate Receptor as Potential Imaging Agents for Brain

Abnormal activity of various <i>N</i>-methyl-d-aspartate receptor (NMDAR) subtypes has been implicated in a wide variety of neurological disorders such as Alzheimer’s disease, schizophrenia, and epilepsy. Imaging agents for PET and SPECT that target NMDARs in a subtype-selective fashion may enable better characterization of those disorders and enhance drug development. On the basis of a pyrazoline derivative that demonstrated neuroprotective effects in vivo, we synthesized a series of <i>para</i>-substituted analogues and measured their affinities to various NMDAR subtypes. Compounds <b>4a</b>–<b>c</b> and <b>4e</b> showed greater, nanomolar affinity for the GluN1/2A subtype versus GluN1/2B. Dicarbomethoxy (pro-drug) analogues of [^124/125I]<b>4d</b> and [¹¹C]<b>4e</b> (i.e., [^124/125I]<b>11d</b> and [¹¹C]<b>11e</b>) were generated and tested for NMDAR binding specificity in ex vivo autoradiography and brain biodistribution studies. Although NMDAR-specific binding could be demonstrated for [¹²⁵I]<b>11d</b> and [¹¹C]<b>11e</b> through autoradiography and biodistribution studies, imaging of neither [¹²⁴I]<b>11d</b> nor [¹¹C]<b>11e</b> could demonstrate brain penetration sufficient for detection by PET