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Liposomes, Exosomes, and Virosomes: From Modeling Complex
Membrane Processes to Medical Diagnostics and Drug Delivery
Poster Abstracts
114
60-POS
Board 30
In Vitro Folding of the Membrane Transporter Leut in Detergent and Lipid Bilayer
Environments
Michael Sanders
, Heather Findlay, Paula Booth.
King's College, London, United Kingdom.
Little is understood about the process that governs the folding of multi domain α-helical integral
membrane proteins from an amino acid sequence to a three dimensional functional structure[1].
The neurotransmitter sodium symporter (NSS) protein family is an example of α-helical
transporter proteins. The NSS family encompasses a wide range of prokaryotic and eukaryotic
ion-coupled transporters that regulate the transport of neurotransmitter molecules whose
dysfunction has been implicated in several neurological and psychiatric disorders [2]. The
prokaryotic orthologue of the NSS family LeuT comes from the organism Aquifex aeolicus.
LeuT is responsible for the uphill transport of Leucine as a part of a symport mechanism driven
by a sodium electrochemical gradient.
In Vitro folding of α-helical membrane proteins has been studied mostly within protein detergent
complexes or unfolded protein has been refolded into a lipid bilayer [3]. Here we exploit
reversible chemical unfolding to determine the unfolding free energy of LeuT both as part of a
protein-detergent complex and within a lipid bilayer. In dodecyl maltoside micelles, LeuT
reversibly unfolds in Urea, whereas in a lipid bilayer LeuT only reversibly unfolds in the
presence of both Urea and the nonionic detergent Octyl-β-Glucoside without physically changing
the state of the lipid bilayer. Chemical denaturation causes a significant reduction in the α-helical
content and loss of transport activity, full recovery of helical content and substrate transport
occurs when the denaturants are removed allowing the free energy to be determined.
This study gives insight into the comparative free energy landscape of protein folding between
detergent and lipid environments as well as the effect of protein-lipid interactions on membrane
protein function and stability.