Distinct recognition of substrates by the human and Drosophila serotonin transporters

The human and Drosophila serotonin transporters (hSERT and dSERT, respectively) were used to explore differences in substrate properties. hSERT and dSERT showed similar Km values for 5-hydroxytryptamine (5-HT; serotonin) transport (1.2 and 0.9 micro M, respectively), suggesting similar recognition of 5-HT by the two species variants. Although dSERT cell surface expression was approximately 8-fold lower than that of hSERT, dSERT does appear to have a 2-fold faster turnover number for inward transport of 5-HT. Interestingly, another substrate, N-methyl-4-phenylpyridinium (MPP+), was transported only by hSERT. However, MPP+ inhibited 5-HT uptake in both species variants with similar potencies. Two cross-species chimeras, H1-118D119-627 and H1-281D282-476H477-638, were also unable to transport MPP+, implicating the role of transmembrane domains V to IX in the substrate permeation pathway. Based on exchange experiments, certain substituted-amphetamines also appear to be poor substrates at dSERT. Two-electrode voltage-clamp studies in oocytes confirmed that the amphetamines do not possess substrate-like properties for dSERT. Our data suggest distinct molecular recognition among SERT substrate classes that influence translocation mechanisms.