Project Details
Projekt
Exploring the structural and mechanistic basis of H+, Na+, and K+ coupling in secondary active glutamate transporters
Applicant
Professorin Dr. Inga Hänelt
Subject Area
Biochemistry
Term
from 2011 to 2012
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 199883430
Ion coupling is a fundamental property of secondary active transporters. The structural and mechanistic basis of ion coupling is not well understood, and is the main topic of this proposal. Glutamate transporters are exquisitely suited to study ion coupling for two reasons. First, crystal structures of the archaeal transporter Gltph are known and provide a framework for the exploration. Second, different members of the glutamate transporter family use different coupling ions, and substrate:coupling ion stoichiometries. The diversity will facilitate the determination of the structural basis of sodium ion, proton and potassium ion selectivity. A structure-guided conversion of the Na+ coupled transporter Gltph into H+ and K+ coupled proteins will be pursued. First large domains will be exchanged between transporters with different ion selectivity, and subsequently the sequences of the chimeras will be fine-tuned. The work will lead to the unraveling of the sequence determinants for ion coupling. In parallel, the chimeric proteins will be included in crystallization experiments, which will provide the structural basis of ion coupling. A side project will concentrate on the time resolved mechanisms of sodium and aspartate binding in Gltph. By combining EPR, single molecule FRET and stopped flow measurements with different crystallization trial the transport cycle will be explained and an improved functional model will result. In summary, the work will provide basic insights in the medically important family of glutamate transporters, and more specifically in the structural basis of cation coupling in secondary transporters.
DFG Programme
Research Fellowships
International Connection
Netherlands
