Systemic lupus erythematosus (SLE) is a multisystem autoimmune disorder with variable clinical expression and incompletely understood causes. Serum bilirubin has been reported to correlate inversely with SLE disease activity and may exert antioxidant and immunomodulatory effects. Spirulina (Arthrospira platensis) contains the phycobiliprotein phycocyanin, whose chromophore phycocyanobilin is an open-chain tetrapyrrole structurally related to bile pigments. Given that reduced serum bilirubin is associated with higher SLE activity and that bilirubin has immunoregulatory properties, the authors investigated whether phycocyanobilin might interact with proteins implicated in SLE pathobiology.
Previous network and similarity analyses identified multiple Spirulina-derived compounds potentially relevant to immune modulation. In the present work the authors focused on phycocyanobilin after structural-similarity screening against bilirubin and performed molecular docking and molecular dynamics (MD) simulations to examine possible molecular interactions with candidate protein targets.
Structural similarity screening
The structural similarity between bilirubin (PubChem CID 5280352) and 833 bioactive compounds identified from A. platensis C1 was evaluated using the PubChem structural similarity tool. Phycocyanobilin was found to share a high Tanimoto similarity score (93%) with bilirubin and was selected for further molecular investigation.
Molecular docking
Molecular docking was carried out using MGLTools with AutoDock. The 3D ligand structures were retrieved from PubChem and receptor structures from the RCSB PDB. Docking targets included several proteins previously implicated in the authors' prior analyses: EGFR (with defined W2R and SO4 binding sites), FYN, HLA-B, LCK, LYN, and TP53. Grid boxes were configured to cover relevant binding domains for each receptor. The docking protocol was validated by re-docking native ligands from crystal structures and calculating RMSD between predicted and experimental poses to confirm the method's reliability.
Target prediction and selection for MD
NetPredictor, a network-based bioinformatics tool, was used to prioritize potential protein receptors for the Spirulina-derived compounds. Based on docking outcomes and NetPredictor results, LYN kinase emerged as the most promising receptor for detailed MD simulation.
Molecular dynamics simulations and analyses
The LYN kinase structure (PDB ID: 3A4O) was prepared by removing waters and heteroatoms. Protonation states were assigned at pH 7.0 using PROPKA 3.0. Energy minimization and MD simulations were performed in AMBER 16 with the ff14SB force field. Simulations ran for 60 ns at 300 K and 1 atm with SHAKE constraints applied to bonds involving hydrogen; the ff14SB force field was used for protein and ligand. System stability was assessed via RMSD for the protein backbone, the complex, and the ligand using PTRAJ. The final 10 ns of MD trajectories were used for binding free energy calculations (MM/PBSA and MM/GBSA), hydrogen bond analysis, and residue-level decomposition to identify key contributors to binding.
Comparative analyses were conducted between the algal-derived ligand–LYN complex and the corresponding native ligand (staurosporine)–LYN complex to contextualize interaction patterns and binding stability.
Structural similarity and docking highlights
Using the PubChem similarity tool, phycocyanobilin displayed a Tanimoto score of 93% relative to bilirubin. Molecular docking across the selected receptors identified favorable predicted binding affinities for phycocyanobilin and bilirubin toward several targets, including EGFR, FYN, HLA-B, LCK, LYN, and TP53. Target prediction using NetPredictor prioritized LYN kinase for further study based on the combined docking and network-analysis results.
MD simulation outcomes for the phycocyanobilin–LYN complex
MD simulations of the phycocyanobilin–LYN complex (60 ns) indicated a stable ligand–protein complex across the production run, with system stability monitored by RMSD metrics for backbone, complex, and ligand. The final 10 ns of trajectories were analyzed in detail. Hydrogen bond analyses and residue decomposition identified specific amino acids contributing favorably to complex stability; these residue-level contributions were reported as part of the MM/PBSA and MM/GBSA decomposition analyses.
Comparison with native ligand (staurosporine)
When compared with the native ligand staurosporine in complex with LYN, the phycocyanobilin–LYN complex demonstrated interaction profiles considered comparable in terms of key contacts and overall stability. Binding free energy calculations using MM/PBSA and MM/GBSA supported strong and stable binding of phycocyanobilin to LYN, with residue-level analyses highlighting contributions from specific interacting residues to the total binding energy. Exact numerical values for binding free energies and per-residue energy contributions are provided in the manuscript and supporting information.
The combined in silico findings suggest that phycocyanobilin can form a stable complex with LYN kinase, a tyrosine kinase implicated in immune-cell signaling pathways relevant to SLE. The high structural similarity to bilirubin motivated the hypothesis that phycocyanobilin might mimic or modulate bilirubin-associated molecular interactions; docking, target prediction, and MD simulation results are consistent with this hypothesis for LYN.
MD-based binding free energy and residue decomposition analyses indicate that multiple interaction types—including van der Waals and electrostatic contacts and specific hydrogen bonds—contribute to complex stability. The authors emphasize that interactions observed in computational models warrant experimental validation to determine whether phycocyanobilin modulates LYN activity in biological systems and whether such modulation could influence SLE-related signaling pathways.
Limitations and next steps
The study is computational and does not include biochemical, cellular, or in vivo validation. Details such as absolute binding affinities in experimental systems, functional effects on LYN kinase activity, and downstream immunological consequences remain to be established. The authors recommend follow-up experimental work to test the functional relevance of the predicted phycocyanobilin–LYN interaction in the context of SLE.
Phycocyanobilin from A. platensis shares strong structural similarity with bilirubin and, in silico, binds stably to LYN kinase with interaction patterns comparable to a native kinase inhibitor (staurosporine). These computational results suggest that phycocyanobilin may modulate LYN-associated signaling pathways and merit further experimental investigation as a potential immunomodulatory bioactive compound relevant to SLE.
All relevant data are reported within the manuscript and its supporting information files, as stated by the authors. The work received no specific funding, and the authors declared no competing interests. Specific numeric results, tables, figures, and methodological parameters referenced in this summary are available in the full PLOS ONE article and its supporting materials.