Monoamine neurotransmitters, including norepinephrine, adrenaline, dopamine, serotonin, and histamine, play important physiological roles in the nervous system and other tissues. Vesicle monoamine transporter 2 (VMAT2) is the only transporter protein in the central nervous system that mediates the storage of monoamine neurotransmitters. VMAT2 utilizes the electrochemical potential of protons to transport neurotransmitters or neurotoxins MPP+into vesicles in a 2:1 stoichiometric ratio of protons to substrates, playing a crucial role in mediating nerve impulse transmission and neuroprotection.

In the early stage, researchers proposed insights into the structure and function of VMAT2, but the mechanism of action of how VMAT2 recognizes and transports other monoamine neurotransmitters with different chemical structures and the Parkinson's disease inducer MPP+is still unclear. Furthermore, the molecular mechanism of proton coupled substrate transport still needs further investigation.

On May 22, Zhao Yan's team from the Institute of Biophysics of the Chinese Academy of Sciences, together with Jiang Daohua's team from the Institute of Physics of the Chinese Academy of Sciences/National Research Center for Condensed Matter Physics in Beijing, published a research paper entitled Structural snapshots of human VMAT2 recent insights into abstract recognition and proton coupling mechanism online on Cell Research. This study reported the apo structure of human vesicular monoamine transporter 2 (hVMAT2) at low pH, which combines vesicular cavity oriented complex structures with norepinephrine, dopamine, histamine, and neurotoxin MPP+, as well as cytoplasmic oriented complex structures with norepinephrine, revealing the structural basis for VMAT2 to recognize different substrates and further improving the molecular mechanism of proton coupled substrate transport.

Although the molecular structures of norepinephrine, dopamine, serotonin, and MPP+vary, the structure of VMAT2 complexes that bind to these substrates indicates that they all bind to similar positions in transport proteins. However, the subtle differences in certain functional groups of these substrate molecules result in differences in their interactions and binding patterns with transporters. This reveals how VMAT2 effectively recognizes different substrate molecules. At the same time, histamine binds at a location different from the binding of other substrates. Based on functional experiments, the researchers validated the reliability of this site.

In addition, the study reported the structures of norepinephrine binding to VMAT2 in different conformations, including cytoplasmic and vesicular orientations. During the conformational changes accompanying substrate transport, The N-terminal domain undergoes approximately 30 ° rotation relative to the C-terminal domain, alternately exposing the substrate binding sites on the cytoplasmic or vesicular side, achieving substrate enrichment in the vesicles. Although VMAT2 underwent significant conformational changes, the substrate binding pocket of norepinephrine remained relatively stable during this process, and the interaction with surrounding residues did not undergo significant changes. This clever conformational transformation feature is crucial for the functionality of VMAT2.

Previous studies have reported that the D399 residue may be a key site for substrate binding and protonation, and a hydrogen bonding network has been discovered within the N-terminal domain. This hydrogen bonding network may play a crucial role in coupling protons and promoting the conformational transition of VMAT2. This study confirmed that D33 may be another key protonation site by analyzing the structure of VMAT2 under different pH conditions. Specifically, Protonation of D33 can induce D33 The local conformational changes in the side chains of R189 and Q192 residues disrupted the hydrogen bonding network, thereby promoting the transition of VMAT2 conformation from vesicular orientation to cytoplasmic orientation.

This study analyzed the mechanism by which VMAT2 recognizes monoamine neurotransmitters and neurotoxin molecules with different chemical structures, proposed a molecular mechanism model for conformational changes in VMAT2, and improved the specific mechanism of proton coupled substrate transport. The above achievements provide useful insights into the transport mode of VMAT2, enrich scientists' understanding of the main co transporter superfamily in transporting substrate molecules, and lay the foundation for drug development and optimization.

The research work was supported by the National Natural Science Foundation of China and the Chinese Academy of Sciences strategic leading science and technology project (Category B). The data collection work of cryo electron microscopy has received technical support from the Bioimaging Center of the Protein Science Research Platform of the Institute of Biophysics, the cryo electron microscopy center and soft matter public instrument platform of the Institute of Physics, and the Biological Microscopic Structure Research Platform of the Institute of Modern Agriculture of Peking University. The functional experiment was assisted by the Institute of Biophysics and the Radioisotope Laboratory of the Institute of Microbiology, Chinese Academy of Sciences.

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The structure of VMAT2 binding neurotransmitters and neurotoxins