Exploring fast proton transfer events associated with lateral proton diffusion on the surface of membranes

Abstract

Proton diffusion (PD) across biological membranes is a fundamental process in many biological systems, and much experimental and theoretical effort has been employed for deciphering it. Here, we report on a spectroscopic probe, which can be tightly tethered to the membrane, for following fast (nanosecond) proton transfer events on the surface of membranes. Our probe is composed of a photoacid that serves as our light-induced proton source for the initiation of the PD process. We use our probe to follow PD, and its pH dependence, on the surface of lipid vesicles composed of a zwitterionic headgroup, a negative headgroup, a headgroup that is composed only from the negative phosphate group, or a positive headgroup without the phosphate group. We reveal that the PD kinetic parameters are highly sensitive to the nature of the lipid headgroup, ranging from a fast lateral diffusion at some membranes to the escape of protons from surface to bulk (and vice versa) at others. By referring to existing theoretical models for membrane PD, we found that while some of our results confirm the quasi-equilibrium model, other results are in line with the nonequilibrium model.

Keywords: excited-state proton transfer; lipid vesicles; molecular dynamics; photoacid; proton diffusion.

Scheme 1.

ESPT photoprotolytic cycle of photoacids. hν, photon absorption; rad, radiative decay.

Scheme 2.

Molecular scheme of C₁₂-HPTS incorporated into a lipid vesicle.

Fig. 1.

(Top) Snapshots from atomistic MD trajectories showing the position of the C₁₂-HPTS dye in the upper and lower leaflets of the three phospholipid bilayers (water molecules are shown as cyan sticks). (Bottom) Density distributions along the bilayer normal of the water (blue), lipids (red), phosphorus atoms of the lipid headgroups (yellow), C₁₂-HPTS pyrene moieties (violet), and C₁₂-HPTS hydroxy groups (green).

Fig. 2.

Absorption spectrum of C₁₂-HPTS incorporated in the different lipid vesicles at different pH values. (Insets) pK_a calculations and the molecular schemes of the lipid. O.D, optical density.

Fig. 3.

Emission spectrum of C₁₂-HPTS incorporated in the different lipid vesicles at different pH values, below the pK_a of the photoacid. (A) Spectrum for each of the lipid vesicles as a function of pH. (B) Spectrum at each of the pH values as a function of the lipid vesicle. The dashed lines in B represent the emission spectrum of HPTS.

Fig. 4.

Time-resolved fluorescence decay of the ROH* form in the four lipid vesicles as a function of pH value. (Insets) Zoomed-in views of the first nanoseconds. IRF, instrument response function.

Fig. 5.

Time-resolved fluorescence decay of the ROH* form at each of the pH values as a function of the lipid vesicle, in comparison to the decay of HPTS. (Insets) Zoomed-in views of the first nanoseconds. IRF, instrument response function.