Computational modeling of the N-terminus of the human dopamine transporter and its interaction with PIP2-containing membranes

• Published in: Proteins, structure, function, and bioinformatics structure, function, and bioinformatics, 2015-05, Vol.83 (5), p.952-969
• Main Authors: Khelashvili, George, Doktorova, Milka, Sahai, Michelle A., Johner, Niklaus, Shi, Lei, Weinstein, Harel
• Format: Article
• Language: English
• Subjects: amphetamine; Cell Membrane - chemistry; Dopamine Plasma Membrane Transport Proteins - chemistry; efflux; electrostatics; Humans; Hydrophobic and Hydrophilic Interactions; Lipid Bilayers - chemistry; molecular dynamics; Molecular Dynamics Simulation; neurotransmitter transporter; Phosphatidylinositol 4,5-Diphosphate - chemistry; Phosphatidylserines - chemistry; phosphorylation; Protein Structure, Secondary; Protein Structure, Tertiary; signaling; sodium symporter
• ISSN: 0887-3585; 1097-0134
• DOI: 10.1002/prot.24792

ABSTRACT The dopamine transporter (DAT) is a transmembrane protein belonging to the family of neurotransmitter:sodium symporters (NSS). Members of the NSS are responsible for the clearance of neurotransmitters from the synaptic cleft, and for their translocation back into the presynaptic nerve terminal. The DAT contains long intracellular N‐ and C‐terminal domains that are strongly implicated in the transporter function. The N‐terminus (N‐term), in particular, regulates the reverse transport (efflux) of the substrate through DAT. Currently, the molecular mechanisms of the efflux remain elusive in large part due to lack of structural information on the N‐terminal segment. Here we report a computational model of the N‐term of the human DAT (hDAT), obtained through an ab initio structure prediction, in combination with extensive atomistic molecular dynamics (MD) simulations in the context of a lipid membrane. Our analysis reveals that whereas the N‐term is a highly dynamic domain, it contains secondary structure elements that remain stable in the long MD trajectories of interactions with the bilayer (totaling >2.2 μs). Combining MD simulations with continuum mean‐field modeling we found that the N‐term engages with lipid membranes through electrostatic interactions with the charged lipids PIP2 (phosphatidylinositol 4,5‐Biphosphate) or PS (phosphatidylserine) that are present in these bilayers. We identify specific motifs along the N‐term implicated in such interactions and show that differential modes of N‐term/membrane association result in differential positioning of the structured segments on the membrane surface. These results will inform future structure‐based studies that will elucidate the mechanistic role of the N‐term in DAT function. Proteins 2015; 83:952–969. © 2015 Wiley Periodicals, Inc.