Surface-localized glycine transporters 1 and 2 function as monomeric proteins in Xenopus oocytes

Na+/Cl--dependent neurotransmitter transporters form a superfamily of trans membrane proteins that share 12 membrane-spanning regions. To gain informat ion about the quaternary structure of these transporter proteins, we hetero logously expressed the glial glycine transporter GlyT1 and its neuronal hom olog GlyT2 in Xenopus oocytes. By using metabolic labeling with [S-35]methi onine or surface labeling with a plasma membrane impermeable reagent follow ed by affinity purification, we separately analyzed the total cellular pool s of newly synthesized GlyTs and its functional plasma membrane-bound fract ions. Upon blue native gel electrophoresis, the surface-localized transport er proteins were found to exist exclusively in complex-glycosylated monomer ic form, whereas a significant fraction of the intracellular GlyT1 and GlyT 2 was core-glycosylated and oligomeric. In contrast, even after treatment w ith the crosslinker glutaraldehyde, surface GlyTs failed to migrate as olig omeric proteins. These results indicate that plasma membrane-bound GlyT1 an d GlyT2 are monomeric proteins, Thus, Na+/Cl--dependent neurotransmitter tr ansporters do not require oligomerization for substrate translocation.