Abstract
Cytosolic phospholipase A 2 (cPLA 2 ) associates with membranes where it hydrolyzes phospholipids containing arachidonic acid to initiate an inflammatory cascade. All-atom molecular dynamics simulations were employed to understand the activation process when cPLA 2 associates with the endoplasmic reticulum (ER) membrane of macrophages where it acts. We found that membrane association causes the lid region of cPLA 2 to undergo a closed-to-open state transition that is accompanied by the sideways movement of loop 495-540, allowing the exposure of a cluster of lysine residues (K488, K541, K543, and K544), which binds the allosteric activator PIP 2 in the membrane. The active site of the open form of cPLA 2 , containing the catalytic dyad residues S228 and D549, exhibited a three-fold larger cavity than the closed form of cPLA 2 in aqueous solution. These findings provide mechanistic insight as to how cPLA 2 ER membrane association promotes major transitions between conformational states critical to allosteric activation and enzymatic phospholipid hydrolysis.
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Abstract
Cytosolic phospholipase A2 (cPLA2) associates with membranes where it hydrolyzes phospholipids containing arachidonic acid to initiate an inflammatory cascade. All-atom molecular dynamics simulations were employed to understand the activation process when cPLA2 associates with the endoplasmic reticulum (ER) membrane of macrophages where it acts. We found that membrane association causes the lid region of cPLA2 to undergo a closed-to-open state transition that is accompanied by the sideways movement of loop 495-540, allowing the exposure of a cluster of lysine residues (K488, K541, K543, and K544), which binds the allosteric activator PIP2 in the membrane. The active site of the open form of cPLA2, containing the catalytic dyad residues S228 and D549, exhibited a three-fold larger cavity than the closed form of cPLA2 in aqueous solution. These findings provide mechanistic insight as to how cPLA2 ER membrane association promotes major transitions between conformational states critical to allosteric activation and enzymatic phospholipid hydrolysis.
Competing Interest Statement
The authors have declared no competing interest.
Abbreviations
- AA
- arachidonic acid
- cPLA2
- Group IVA (GIVA) cytosolic phospholipase A2
- ER
- Endoplasmic reticulum
- HDX-MS
- Hydrogen/Deuterium exchange-mass spectrometry
- MD
- molecular dynamics
- PAPC
- 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine
- PC
- phosphatidylcholine
- PCA
- principal component analysis
- PLA2
- Phospholipase A2
- PI
- phosphatidylinositol
- PIP2
- phosphatidylinositol (4,5)-bisphosphate
- PS
- phosphatidylserine
- RMSF
- Root-mean-square fluctuation
- RMSD
- Root-mean-square deviation
- SAPI
- 1-stearoyl-2-arachidonoyl phosphatidylinositol
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