Structure, dynamics, and ion conductance of the phospholamban pentamer

Christopher Maffeo, and Aleksei Aksimentiev
Biophys J 96(12) 4853-65 (2009)
DOI:10.1016/j.bpj.2009.03.053  PMID:19527644  BibTex

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Muscle cells respond to external nerve stimuli by releasing Ca2+ signaling ions from a storage organelle called the sarcoplasmic reticulum (SR). The Ca2+ exposes myosin binding sites on actin filaments, which allows myosin to ratchet along actin and causes the cell to contract. In order for muscle fibers to relax, Ca2+ must be transported from the cytoplasm back to the SR. The Ca2+-ATPase resides in the membrane of the SR and transports two Ca2+ ions against a concentration gradient by using energy of ATP hydrolysis. The uptake of Ca2+ by Ca2+-ATPase is regulated by the membrane protein phospholamban (PLN), which acts as an inhibitor. Although more than 75% of PLN in a lipid bilayer membrane is pentameric, the only known function of PLN, i.e. Ca2+-ATPase inhibition, involves a PLN monomer and not a pentamer. Using all-atom and coarse grained molecular dynamics simulations, we investigated structural dynamics and conductance properties of the recently reported NMR structure of the PLN pentamer. Our simulations demonstrated that, in a lipid bilayer membrane or a detergent micelle, the cytopasmic part of the pentamer undergoes large structural fluctuations while the transmembrane part of the pentamer shrinks and becomes asymmetric. Bound states between neighboring cytoplasmic helices were observed suggesting that the cytoplasmic region may facilitate recognition between oligomerizing protomers. Using steered molecular dynamics simulations, we investigated the feasibility of ion conductance through the pore of a PLN pentamer. The resulting approximate potentials of mean force indicate that the PLN pentamer is unlikely to function as an ion channel. This work is described in a report that appeared in Biophysical Journal.

Abstract

A 52-residue membrane protein, phospholamban (PLN) is an inhibitor of an adenosine-5'-triphosphate-driven calcium pump, the Ca2+-ATPase. Although the inhibition of Ca2+-ATPase involves PLN monomers, in a lipid bilayer membrane, PLN monomers form stable pentamers of unknown biological function. The recent NMR structure of a PLN pentamer depicts cytoplasmic helices extending normal to the bilayer in what is known as the bellflower conformation. The structure shows transmembrane helices forming a hydrophobic pore 4 Å in diameter, which is reminiscent of earlier reports of possible ion conductance through PLN pentamers. However, recent FRET measurements suggested an alternative structure for the PLN pentamer, known as the pinwheel model, which features a narrower transmembrane pore and cytoplasmic helices that lie against the bilayer. Here, we report on structural dynamics and conductance properties of the PLN pentamers from all-atom (AA) and coarse-grained (CG) molecular dynamics simulations. Our AA simulations of the bellflower model demonstrate that in a lipid bilayer membrane or a detergent micelle, the cytoplasmic helices undergo large structural fluctuations, whereas the transmembrane pore shrinks and becomes asymmetric. Similar asymmetry of the transmembrane region was observed in the AA simulations of the pinwheel model; the cytoplasmic helices remained in contact with the bilayer. Using the CG approach, structural dynamics of both models were investigated on a microsecond timescale. The cytoplasmic helices of the CG bellflower model were observed to fall against the bilayer, whereas in the CG pinwheel model the conformation of the cytoplasmic helices remained stable. Using steered molecular dynamics simulations, we investigated the feasibility of ion conductance through the pore of the bellflower model. The resulting approximate potentials of mean force indicate that the PLN pentamer is unlikely to function as an ion channel.