Ceramide-1-phosphate transfer protein enhances lipid transport by disrupting hydrophobic lipid–membrane contacts

Abstract

Cellular distributions of the sphingolipid ceramide-1-phosphate (C1P) impact essential biological processes. C1P levels are spatiotemporally regulated by ceramide-1-phosphate transfer protein (CPTP), which efficiently shuttles C1P between organelle membranes. Yet, how CPTP rapidly extracts and inserts C1P into a membrane remains unknown. Here, we devise a multiscale simulation approach to elucidate biophysical details of CPTP-mediated C1P transport. We find that CPTP binds a membrane poised to extract and insert C1P and that membrane binding promotes conformational changes in CPTP that facilitate C1P uptake and release. By significantly disrupting a lipid’s local hydrophobic environment in the membrane, CPTP lowers the activation free energy barrier for passive C1P desorption and enhances C1P extraction from the membrane. Upon uptake of C1P, further conformational changes may aid membrane unbinding in a manner reminiscent of the electrostatic switching mechanism used by other lipid transfer proteins. Insertion of C1P into an acceptor membrane, eased by a decrease in membrane order by CPTP, restarts the transfer cycle. Most notably, we provide molecular evidence for CPTP’s ability to catalyze C1P extraction by breaking hydrophobic C1P–membrane contacts with compensatory hydrophobic lipid–protein contacts. Our work, thus, provides biophysical insights into how CPTP efficiently traffics C1P between membranes to maintain sphingolipid homeostasis and, additionally, presents a simulation method aptly suited for uncovering the catalytic mechanisms of other lipid transfer proteins.

The sterol transporter STARD3 transports sphingosine at ER-lysosome contact sites

“…The tail-first orientation observed in STARD3 for sphingosine’s entry into the hydrophobic cavity is advantageous for lipid release into the target membrane. Recent studies have also confirmed this orientation in other proteins like C1PTP and ORP3 ORD 42–44 , further supporting the idea that this may be a generalizable mechanism for lipid transfer across various protein systems.…”

Model mechanism for lipid uptake by the human STARD2/PC-TP phosphatidylcholine transfer protein

“…In all simulations where tail insertion events were observed, the headgroup of the PC lipid remained in the aromatic cage, and the phosphate group remained hydrogen bonded to residues R78, Y72, and Q157. No spontaneous release is observed unlike what we observed in CERT/STARD11, 11 indicating that the simulations do not sample interactions or events that would lower the energy barrier for the release of the lipid.…”

“…For instance, molecular simulations of the ceramide-1-phosphate transfer protein (CPTP) showed that the uptake and release of ceramide-1-phosphate by CPTP is facilitated by conformational changes of the CPTP coupled with the disruption of lipids packing below the protein, and the creation of protein-cargo hydrophobic contacts. 11 In simulations of the ceramide transfer protein CERT (STARD11) with lipid bilayers we also observed that the opening of the gate is concomitant to changes in lipid tails packing. In addition, the intercalation of a single phosphatidylcholine lipid in the cavity disrupts the LTP-cargo interactions, and facilitates the release of the ceramide cargo through the polar membrane interface.…”

A Membrane-Assisted Mechanism for the Release of Ceramide from the CERT START Domain

“…This results in looser lipid packing, increased mobility of the lipid tails toward the open hydrophobic cavity, and snorkeling, as well as increased interfacial hydrophobicity as also shown in other studies. 69,70,75 The Journal of Physical Chemistry B Interestingly, the binding and opening mechanisms of the START domain do not appear to be influenced by lipid composition, suggesting the domain's lipid-insensitive nature. However, the cargo release mechanism indicates sensitivity to the membrane lipid composition, implying a potential role of the membrane in the release of the ceramide.…”

“…This interfacial hydrophobic pool provides a nonpolar environment that shields the hydrophobic lipid tails from contact with water or the polar head groups of the membrane lipids. This results in looser lipid packing, increased mobility of the lipid tails toward the open hydrophobic cavity, and snorkeling, as well as increased interfacial hydrophobicity as also shown in other studies. ,, …”