Ions Modulate Key Interactions between pHLIP and Lipid Membranes

Abstract

The pH-low insertion peptide (pHLIP) is used for targeted delivery of drug cargoes to acidic tissues such as tumors. The extracellular acidosis found in solid tumors triggers pHLIP to transition from a membrane-adsorbed state to fold into a transmembrane a-helix. Different factors influence the acidity required for pHLIP to insert into lipid membranes. One of them is the lipid headgroup composition, which defines the electrostatic profile of the membrane. However, the molecular interactions that drive the adsorption of pHLIP to the bilayer surface are poorly understood. In this study, we combine biophysical experiments and all-atom molecular dynamics simulations to understand the role played by electrostatics in the interaction between pHLIP and a 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine bilayer. We observed that the solution ionic strength affects the structure of pHLIP at the membrane surface as well as the acidity needed for different steps in the membrane insertion process. In particular, our simulations revealed that an increase in ionic strength affected both pHLIP and the bilayer; the coordination of sodium ions with the C-terminus of pHLIP led to localized changes in helicity, whereas the coordination of sodium ions with the phosphate moiety of the phosphocholine headgroups had a condensing effect on our model bilayer. These results are relevant to our understanding of environmental influences on the ability of pHLIP to adsorb to the cell membrane and are useful in our fundamental understanding of the absorption of pH-responsive peptides and cell-penetrating peptides.

Intramolecular interactions play key role in stabilization of pHLIP at acidic conditions

“…In addition, lower pH depresses intramolecular interactions in the interior of pHLIP (i.e., R11, D14, and D25). Consistent with our previous study on pHLIP in state II, 37 physiological salt concentrations screen terminal acidic residues, which in turn can lead to increased interactions between interior charged residues (e.g., R11 with either D14 or D25). We realize that environmental factors contribute to conformational changes in pHLIP, but there are obviously other factors that come into play that have not been addressed in this study.…”

“…This particular modification is very similar in nature to the dual salt bridges that give ribonuclease A a very sensitive response to changes in pH 22 . We have previously observed this behavior with R11–D14 salt bridges in state I 15 and state II 37 …”

“…The first is screening of specific residues in both pHLIP and P20G. Salt at physiological concentrations can screen acidic residues in pHLIP 37 . However, these interactions are nonuniform: E3, D14, and D25 have noticeably lower affinity for Na + cations (Figure S12).…”

Targeting Acidic Diseased Tissues by pH-Triggered Membrane-Associated Peptide Folding

“…pHLIPs containing fewer hydrophobic residues have lower affinities for membranes and much faster blood clearance (Weerakkody et al, 2013;Demoin et al, 2016). In addition, peptide adsorption to the membrane is modulated by membrane composition (Kyrychenko et al, 2015;Karabadzhak et al, 2018;Vasquez-Montes et al, 2018) and the local ionic environment (Schlebach, 2019;Vasquez-Montes et al, 2019;Westerfield et al, 2019). Significant attention has been given to the conformational states of pHLIPs adsorbed by membranes (Brown et al, 2014;Gupta et al, 2018;Vasquez-Montes et al, 2018).…”

Ions at the Interface: Pushing the pK of pHLIP

“…Perhaps unsurprisingly, the organization of phospholipids within the bilayer appears to significantly influence pHLIP's membrane insertion (9). In this issue, Westerfield et al (10) explore the influence of salt on the structural properties of both pHLIP and the membrane interface itself and how these relate to changes in apparent pK. The authors first show that physiological salt concentrations reduce the apparent pK of pHLIP.…”