Structural basis of norepinephrine recognition and transport inhibition in neurotransmitter transporters

Abstract

Norepinephrine is a biogenic amine neurotransmitter that has widespread effects on alertness, arousal and pain sensation. Consequently, blockers of norepinephrine uptake have served as vital tools to treat depression and chronic pain. Here, we employ the Drosophila melanogaster dopamine transporter as a surrogate for the norepinephrine transporter and determine X-ray structures of the transporter in its substrate-free and norepinephrine-bound forms. We also report structures of the transporter in complex with inhibitors of chronic pain including duloxetine, milnacipran and a synthetic opioid, tramadol. When compared to dopamine, we observe that norepinephrine binds in a different pose, in the vicinity of subsite C within the primary binding site. Our experiments reveal that this region is the binding site for chronic pain inhibitors and a determinant for norepinephrine-specific reuptake inhibition, thereby providing a paradigm for the design of specific inhibitors for catecholamine neurotransmitter transporters.

Molecular Imaging-Derived Biomarker of Cardiac Nerve Integrity — Introducing High NET Affinity PET Probe 18F-AF78

“…In addition to the in vitro and in vivo studies, an in silico hypothesis might hint at a potential explanation for the high NET affinity of AF78. Because there is currently no available X-ray structure of NET, PDB6M0Z, a dopamine transporter with NET-like mutations (D121G/S426 M/F471 L) in NE-bound form, was first chosen for the docking study 34 . Adopting the conformation of PBD6M0Z, the 3-dimensional structure of NET as a Homo-AF model was established using AlphaFold prediction.…”

“…Adopting the conformation of PBD6M0Z, the 3-dimensional structure of NET as a Homo-AF model was established using AlphaFold prediction. In this model, the smaller radiotracers, including HED, MIBG, 4F-MHPG, 3F-PHPG and 6F-DA ( Table 1 ), can be docked into the primary binding site in a pattern similar to NE, with amino or guanidine groups that fit into the binding pocket A, while those radiotracers with a hydroxy group may form a hydrogen bond with A145 according to the model established by Pidathala ( Figure 6 ) 34 . However, the molecules with a “tail” (3-fluoropropyl group, e.g., FBBG and AF78) do not fit well into the proposed binding pockets as the “tail” flips between binding pocket B and C ( Figure 6 A ).…”

Functional characterization of dopamine and norepinephrine transport across the apical and basal plasma membranes of the human placental syncytiotrophoblast

“…However, the sensitivity of the uptake to selective SERT (paroxetine) and NET (nisoxetine) inhibitors suggests an overlapping substrate specificity whereby both SERT and NET contribute to placental uptake of dopamine across the maternal-facing membrane. Indeed, accumulating evidence reveals that the SLC6A members share structural and mechanistic properties 47 , 48 . Consequently, dopamine has already been described as an alternative substrate of SERT and NET in neuronal tissue, albeit with a mechanistically distinct mode of substrate translocation 49 , 50 .…”

Phenotype-Based HPLC-Q-TOF-MS/MS Coupled With Zebrafish Behavior Trajectory Analysis System for the Identification of the Antidepressant Components in Methanol Extract of Anshen Buxin Six Pills

“…The knockout of SERT and NET reduced immobility in forced swimming test and tail suspension test ( Kalueff et al, 2007 ; Solich et al, 2020 ). Chronic stress leads to the increase of content of SERT and NET, which may be the reason for the decrease of NE and SERT level in brain tissue ( Pidathala et al, 2021 ). Therefore, regulating the expression of SERT and NET plays an important role in the occurrence of depression.…”