Secreted phospholipase A2s (sPLA2s) participate in physiological function by their enzyme and receptor binding activity. Muscle-type phospholipase A2 receptor (M-type PLA2R) is the sPLA2 binding protein with the highest affinity so far, and also inhibits the enzyme activity of sPLA2. There is species specificity and pH dependence for the binding of M-type PLA2R to sPLA2. Mouse sPLA2 Group IIE (mGIIE) has been verified to have a high affinity for mouse M-type PLA2R (M-type mPLA2R) at the nanomolar scale. For further exploration of the receptor binding mechanism of GIIE, in this study, we use Alphafold Multimer to generate complex models of mGIIE with the M-type mPLA2R ectodomain, wild-type CTLD5 domain of mPLA2R, and three CTLD5 mutants, respectively. mPLA2R-mGIIE models exhibit heterogeneous extended mPLA2R conformations with uncovered sPLA2-binding surface of CTLD5 domain. Complexed models of mGIIE with wild-type and mutated mCTLD5 further confirm that helix α1 of mCTLD5, especially essential residues F838 and W842, interact with the substrate pocket of mGIIE and thus inhibit its enzyme activity. Peptides from helix α1 of mCTLD5 are verified to inhibit the enzymatic activity of mGIIE. This AI-guided research would substantially accelerate our understanding of the functional study of GIIE, and provide the lead-peptide for the further inhibitor design of sPLA2.
This study aimed to design a novel liposome containing GA modified phosphatidylcholine lipid (GA-PC Lip) and determine its susceptibility to tumor over-expressed secretory phospholipase A2 (sPLA2) and its anti-cancer effect compared to conventional liposomes (Convention Lip). The liposomes were characterized for size, drug loading, encapsulation efficiency, and stability. A 6-CF release assay was conducted to assess the sensitivity of the liposomes to the tumor-overexpressed secretory phospholipase A2 (sPLA2). In vitro experiment, the sPLA2 levels in the Colo205 cell culture medium were detected by the Elisa kit and the anti-cancer effect of the oxaliplatin (L-OHP) loaded GA-PA Lip was analyzed by the CCK-8 assay. Results showed that both of L-OHP loaded formulations (GA-PC Lip and Convention Lip) had similar particle sizes of ∼100 nm and close entrapment efficiency values of 4.5–4.8%. The results of CF release assay indicated that the labeled GA-PC Lip had released more quickly than CF labeled Convention Lip in the presence of Bv sPLA2 and GA-PC Lip had a release of about 95% 6-CF at 2 h, whereas Convention Lip only released about 13% 6-CF. In addition, the average concentrations of sPLA2 in the cell-conditioned medium (CCM) of Colo205 cancer cells increased with incubation time and L-OHP loaded GA-PC Lip had much greater anti-proliferative activity than Convention Lip against Colo205 cells. These findings suggest that GA-PC Lip is an ideal complex for sPLA2-triggered release and has potential applications in enzyme-triggered smart anti-cancer drug release system to increase the anti-cancer effect.
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