Coupled Electrostatic and Hydrophobic Destabilisation of the Gelsolin-Actin Complex Enables Facile Detection of Ovarian Cancer Biomarker Lysophosphatidic Acid

Abstract

Lysophosphatidic acid (LPA) is a promising biomarker candidate to screen for ovarian cancer (OC) and potentially stratify and treat patients according to disease stage. LPA is known to target the actin-binding protein gelsolin which is a key regulator of actin filament assembly. Previous studies have shown that the phosphate headgroup of LPA alone is inadequate to bind to the short chain of amino acids in gelsolin known as the PIP₂-binding domain. Thus, the molecular-level detail of the mechanism of LPA binding is poorly understood. Here, we model LPA binding to the PIP₂-binding domain of gelsolin in the gelsolin-actin complex through extensive ten-microsecond atomistic molecular dynamics (MD) simulations. We predict that LPA binding causes a local conformational rearrangement due to LPA interactions with both gelsolin and actin residues. These conformational changes are a result of the amphipathic nature of LPA, where the anionic phosphate, polar glycerol and ester groups, and lipophilic aliphatic tail mediate LPA binding via charged electrostatic, hydrogen bonding, and van der Waals interactions. The negatively-charged LPA headgroup binds to the PIP₂-binding domain of gelsolin-actin while its hydrophobic tail is inserted into actin, creating a strong LPA-insertion pocket that weakens the gelsolin-actin interface. The computed structure, dynamics, and energetics of the ternary gelsolin-LPA-actin complex confirms that a quantitative OC assay is possible based on LPA-triggered actin release from the gelsolin-actin complex.

Keywords: actin; gelsolin; lipid-protein interaction; lysophosphatidic acid; ovarian cancer; predictive molecular modelling.

Figure 1

The polar and nonpolar regions of lysophosphatidic acid (LPA above) and phosphatidylinositol 4,5-bisphosphate (PIP₂ below).

Figure 2

(A) The LPA-bound gelsolin(1–3)-actin complex: five numbered Ca²⁺ ions, actin (blue) and gelsolin(1–3) (orange, with the PIP₂-binding domain coloured yellow). The arrow points to the LPA molecule, shown in stick representation, coordinated to Ca²⁺ #3. An ATP molecule binds Ca²⁺ #5. (B) The distinct domains of gelsolin(1–3), from left to right, G1 (violet), G2 (yellow), and G3 (orange).

Figure 3

LPA docked to gelsolin(1–3)-actin in Dock 1 (pink), Dock 2 (green), and Dock 3 (light blue) models. Actin is dark blue, gelsolin is orange, and the PIP₂-binding domain in gelsolin is coloured yellow in both the cartoon (left) and molecular surface representation (right) of the protein complex.

Figure 4

Actin residues within 3 Å of LPA following 2 µs of dynamics, defining the actin LPA-insertion pocket from two different starting structures: (A) Dock 2 and (B) Dock 3. (C) LPA binding to the gelsolin(1–3)-actin complex after 2 µs from the starting structures of Dock 2 (pink) and Dock 3 (light blue), with (D) the inset projecting on the designated LPA-insertion pocket. Residues that are within 3 Å of LPA are coloured based on polarity: nonpolar (white), polar (green), basic (blue), and acidic (red).

Figure 5

Timelines of van der Waals and electrostatic contributions to the interaction energies between (A,B) gelsolin(1–3) and actin and (C,D) the gelsolin(1–3)-actin complex and LPA.

Figure 6

Timelines of binding energies of LPA with gelsolin(1–3)-actin in (A) Dock 2 and (B) Dock 3 models.

Figure 7

Residue-wise decomposition of binding energies in the gelsolin(1–3)-actin-LPA complex identifying the local conformational change of Lys150 in gelsolin(1–3), where (A) Lys150 is H-bonded with actin during early stage sub-200 ns dynamics and (B) Lys150 H-bonds with LPA in the last 200 ns of dynamics.