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IBI SEMINAR
Molecular mechanisms of sodium-coupled glutamate
uptake Peter Larsson Associate
Professor Department of Physiology and
Biophysics University of Miami Tuesday November
24, 2009 1200 Noon Biological Sciences
211 University of Calgary Host Sergei Noskov
Abstract Glutamate transporters remove glutamate
from synapse to maintain an efficient synaptic
transmission and prevent glutamate from reaching
neurotoxic levels. Glutamate transporters couple
the uptake of one glutamate to the co-transport
of three Na and H and the counter-transport of
one K. The molecular mechanism for this coupled
uptake of glutamate and its co- and
counter-transported ions is not known. In a
crystal structure of the bacterial glutamate
transporter homologue, GltPH, only two cations
are bound to the transporter and there was no
indication of the location of the third Na site.
Using Voltage Clamp Fluorometry (VCF) and
computer simulations, we identify the third Na
binding site. Interestingly, the Na bound at
this site forms part of the binding site for the
amino acid substrate, which suggests an
explanation for the strict coupling of Na
transport to uptake of glutamate in glutamate
transporters. Hairpin two (HP2) has been proposed
as the extracellular gate of glutamate
transporters. To test this hypothesis, we use
double site-directed spin-labeling electron
paramagnetic resonance spectroscopy on the
bacterial aspartate transporter GltPh to examine
conformational changes in HP2. Surprisingly, the
two co-ligands Na and aspartate induce opposite
movements of HP2. We find that Na binding to the
transporter opens the extracellular gate, while
the subsequent binding of aspartate closes the
gate. Our findings are consistent with HP2
comprising the extracellular gate of glutamate
transporters and that Na binding opens the
extracellular gate thereby allowing for amino
acid substrate binding.
Pizza and pop will be provided