Structural basis of norepinephrine recognition and transport inhibition in neurotransmitter transporters

Abstract

Highlights• X-ray structures of the Drosophila dopamine transporter in substratefree and norepinephrine bound forms. • Norepinephrine and dopamine bind in distinct conformations within the binding pocket. • Chronic pain inhibitors S-duloxetine, milnacipran and tramadol bind in the primary binding site and overlap with the norepinephrine-binding pose. • Selective norepinephrine reuptake inhibition occurs through specific interactions at the subsite C in the primary binding pocket. AbstractNorepinephrine is a biogenic amine neurotransmitter that has widespread effects on cardiovascular tone, alertness and sensation of pain. As a consequence, blockers of norepinephrine uptake have served as vital tools to treat depression and chronic pain. Here, we employ a modified Drosophila melanogaster dopamine transporter as a surrogate for the human norepinephrine transporter and determine the 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.Keywords. L-Norepinephrine (NE), Drosophila melanogaster dopamine transporter (dDAT), dopamine (DA), serotonin (5-HT), solute carrier family, secondary active transport, neurotransmitter sodium symporters, duloxetine, milnacipran, tramadol.

Assessment of Paroxetine Molecular Interactions with Selected Monoamine and γ-Aminobutyric Acid Transporters

“…It was isolated in 1901 and is the first hormone obtained in the crystalline state [54]. The presence of norepinephrine in two main enantiomers (R/S-NE) was confirmed by structural (crystallographic and spectroscopic studies [54,55]). In crystallographic study it was proved that NE has a zwitterionic structure, formed by proton transfer from the meta phenolic hydroxyl group to the nitrogen atom ( − O(OH)Ph-CH(OH)-CH 2 -NH 3 + ) [56].…”

“…It was one of the first neurotransmitters to be discovered [53]. It was isolated in 1901 and is the first hormone obtained in the crystalline state [54]. The presence of norepinephrine in two main enantiomers (R/S-NE) was confirmed by structural (crystallographic and spectroscopic studies [54,55]).…”