Chimeric vesicular monoamine transporters identify structural domains that influence substrate affinity and sensitivity to tetrabenazine.
The vesicular monoamine transporters (VMATs) 1 and 2 show close sequence similarity but substantial differences in apparent substrate affinity and drug sensitivity. To identify structural domains that determine these functional characteristics, chimeric transporters were constructed and their properties were analyzed in a heterologous expression system. The results implicate multiple regions in the recognition of serotonin and histamine and the sensitivity to tetrabenazine. Two domains of VMAT2, one extending from transmembrane domain (TMD) 5 to the beginning of TMD8 and the other from the end of TMD9 through TMD12, increase the affinity for serotonin and histamine as well as the sensitivity to tetrabenazine but only in the context of more C-terminal and more N-terminal VMAT2 sequences, respectively. In addition, the extreme N terminus of VMAT2 alone suffices to confer a partial increase in substrate affinity and tetrabenazine sensitivity. Despite these similarities among the interactions with serotonin, histamine, and tetrabenazine, the region of VMAT2 from TMD3 through TMD4 increases serotonin affinity but not histamine affinity or tetrabenazine sensitivity, and whereas the region from TMD5 to TMD8 of VMAT2 increases serotonin affinity in the context of more C-terminal VMAT2 sequences, the region encompassing TMD5 through TMD7 reduces serotonin but not histamine affinity or tetrabenazine sensitivity in the context of more N-terminal VMAT2 sequences. Thus, the chimeric analysis also reveals differences between serotonin recognition and the recognition of both histamine and tetrabenazine that may account for the observed differences in their interaction with the transport protein.
Peter
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Department of Neurology, University of California School of Medicine, Los Angeles, California 90024, USA.
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Edwards
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D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002522:Chlorocebus aethiops; D006632:Histamine / Q000378:metabolism; D007700:Kinetics; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism*; D026901:Membrane Transport Proteins; D016297:Mutagenesis, Site-Directed; D009479:Neuropeptides; D018377:Neurotransmitter Agents / Q000378:metabolism; D017354:Point Mutation; D017433:Protein Structure, Secondary; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D012110:Reserpine / Q000494:pharmacology; D012701:Serotonin / Q000378:metabolism; D013379:Substrate Specificity; D013747:Tetrabenazine / Q000378:metabolism / Q000494:pharmacology*; D014162:Transfection; D050492:Vesicular Biogenic Amine Transport Proteins; D050493:Vesicular Monoamine Transport Proteins
D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D009479:Neuropeptides; D018377:Neurotransmitter Agents; D011993:Recombinant Fusion Proteins; C493454:Slc18a2 protein, rat; D050492:Vesicular Biogenic Amine Transport Proteins; D050493:Vesicular Monoamine Transport Proteins; D012701:Serotonin; D006632:Histamine; D012110:Reserpine; D013747:Tetrabenazine