Molecular determinants of transport stimulation of EAAT 2 are located at interface between the trimerization and substrate transport domains

• Published in: Journal of neurochemistry 2015-04, Vol.133 (2), p.199-210
• Main Authors: Mortensen, Ole V., Liberato, José L., Coutinho‐Netto, Joaquim, dos Santos, Wagner F., Fontana, Andréia C. K.
• Format: Article
• Language: English
• ISSN: 0022-3042; 1471-4159
• DOI: 10.1111/jnc.13047

Excitatory amino acid transporters ( EAAT s) regulate glutamatergic signal transmission by clearing extracellular glutamate. Dysfunction of these transporters has been implicated in the pathogenesis of various neurological disorders. Previous studies have shown that venom from the spider Parawixia bistriata and a purified compound (Parawixin1) stimulate EAAT 2 activity and protect retinal tissue from ischemic damage. In the present study, the EAAT 2 subtype specificity of this compound was explored, employing chimeric proteins between EAAT 2 and EAAT 3 transporter subtypes and mutants to characterize the structural region targeted by the compound. This identified a critical residue (Histidine‐71 in EAAT 2 and Serine‐45 in EAAT 3) in transmembrane domain 2 ( TM 2) to be important for the selectivity between EAAT 2 and EAAT 3 and for the activity of the venom. Using the identified residue in TM 2 as a structural anchor, several neighboring amino acids within TM 5 and TM 8 were identified to also be important for the activity of the venom. This structural domain of the transporter lies at the interface of the rigid trimerization domain and the central substrate‐binding transport domain. Our studies suggest that the mechanism of glutamate transport enhancement involves an interaction with the transporter that facilitates the movement of the transport domain. image We identified a domain (purple star) in the glutamate transporter EAAT2 that is important for transport stimulation through a spider venom, and suggest a mechanism for enhanced transporter function through facilitated substrate translocation (arrow). Because the dysfunction of glutamate transporters is implicated in the pathogenesis of neurological disorders, understanding the mechanisms of enhanced transport could have therapeutic implications.