Title

Membrane-Induced p Ka Shifts in wt-pHLIP and Its L16H Variant

Document Type

Article

Date of Original Version

6-12-2018

Abstract

The pH (low) insertion peptides (pHLIPs) is a family of peptides that are able to insert into a lipid bilayer at acidic pH. The molecular mechanism of pHLIPs insertion, folding, and stability in the membrane at low pH is based on multiple protonation events, which are challenging to study at the molecular level. More specifically, the relation between the experimental pK of insertion (pKexp) of pHLIPs and the pKa of the key residues is yet to be clarified. We carried out a computational study, complemented with new experimental data, and established the influence of (de)protonation of titrable residues on the stability of the peptide membrane-inserted state. Constant-pH molecular dynamics simulations were employed to calculate the pKa values of these residues along the membrane normal. In the wt-pHLIP, we identified Asp14 as the key residue for the stability of the membrane-inserted state, and its pKa value is strongly correlated with the experimental pKexp measured in thermodynamics studies. Also, in order to narrow down the pH range at which pHLIP is stable in the membrane, we designed a new pHLIP variant, L16H, where Leu in the 16th position was replaced by a titrable His residue. Our results showed that the L16H variant undergoes two transitions. The calculated pKa and experimentally observed pKexp values are in good agreement. Two distinct pKexp values delimit a pH range where the L16H peptide is stably inserted in the membrane, while, outside this range, the membrane-inserted state is destabilized and the peptide exits from the bilayer. pHLIP peptides have been successfully used to target cancer cells for the delivery of diagnostics and therapeutic agents to acidic tumors. The fine-tuning of the stability of the pHLIP inserted state and its restriction to a narrow well-defined pH range might allow the design of new peptides, able to discriminate between tissues with different extracellular pH values.

Publication Title

Journal of Chemical Theory and Computation

Volume

14

Issue

6

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