Characterization of multiple forms of the human glycine transporter type-2.
The human glycine transporter type 2 (hGlyT2) was cloned from a spinal cord cDNA library using PCR-based methodologies. The isolated sequence exhibits 89% homology with the previously isolated rat GlyT2 cDNA (Liu et al., J. Biol. Chem. 268 (1993) 22802-22808) at the nucleotide level, and 93% amino acid sequence identity. The greatest divergence between the human and rat sequences is found at the amino-terminus, where only 74% amino acid identity exists in residues 1-190. Expression of the intact hGlyT2 transporter sequence in COS-7 cells resulted in a 10-fold increase in high-affinity uptake relative to control cells transfected with vector alone. An artificially truncated form of the transporter, missing the NH2-terminal 153 amino acids, was also capable of mediating glycine uptake. However, an identified variant lacking the first 234 amino acids was non-functional. An hGlyT2 transporter containing a 14-residue deletion in the intracellular loop between transmembrane domains 6 and 7 was also identified and expressed, but failed to mediate glycine uptake. Like rat GlyT2, the high-affinity uptake mediated by hGlyT2 was found to be insensitive to the GlyT1 inhibitor sarcosine.
Gallagher
M J
MJ
Discovery Research, Gliatech, 23420 Commerce Park Rd., Cleveland, OH 44122, USA. gallagherm@gliatech.com
Burgess
L H
LH
Brunden
K R
KR
D020121:5' Untranslated Regions; D000595:Amino Acid Sequence; D026921:Amino Acid Transport Systems, Neutral; D000818:Animals; D001483:Base Sequence; D001692:Biological Transport / Q000187:drug effects; D019556:COS Cells; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D050487:Glycine Plasma Membrane Transport Proteins; D006801:Humans; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D016133:Polymerase Chain Reaction; D020033:Protein Isoforms / Q000037:antagonists & inhibitors / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D012521:Sarcosine / Q000494:pharmacology; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D013116:Spinal Cord / Q000737:chemistry; D014162:Transfection
D020121:5' Untranslated Regions; D026921:Amino Acid Transport Systems, Neutral; D002352:Carrier Proteins; D018076:DNA, Complementary; D050487:Glycine Plasma Membrane Transport Proteins; D009419:Nerve Tissue Proteins; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; C493271:SLC6A5 protein, human; C493268:SLC6A9 protein, human; C493269:Slc6a5 protein, rat; C493266:Slc6a9 protein, rat; D012521:Sarcosine