Decoding the catalytic potential of a Cycloclasticus zancles ring-hydroxylating dioxygenase through computational analysis for enhanced PAH biodegradation.

Polycyclic aromatic hydrocarbons (PAHs) are widespread, toxic, and recalcitrant pollutants in marine ecosystems. Microbial biodegradation offers a sustainable remediation strategy, with Cycloclasticus zancles recognized as a keystone degrader. Here, we used in silico approaches to characterize a ring-hydroxylating dioxygenase (RHD), the enzyme catalyzing the initial and rate-limiting step of PAH catabolism. The AlphaFold-predicted RHD structure was validated using PROCHECK and ERRAT, confirming strong stereochemical and structural reliability. Toxicity screening classified all twenty PAHs as Cramer Class III, underscoring the ecological risk and the need for effective degradation. Protein-protein interaction and co-expression analyses revealed associations with electron transfer partners and conserved dioxygenase-related genes, suggesting coordinated regulation of hydrocarbon metabolism. Molecular docking showed high binding affinities for large PAHs, with benzo[a]pyrene (- 9.3 kcal/mol), benzo[b]fluoranthene (- 9.1 kcal/mol), and benzo[e]pyrene (- 9.0 kcal/mol) forming the most stable complexes. Interaction profiling highlighted stabilization through hydrophobic contacts, π-π stacking, and hydrogen bonding with residues including Phe223, Tyr276, Leu278, Asn252, and Gln309. Collectively, these results demonstrate the enzyme's catalytic adaptability, biosafety relevance, and ecological significance, establishing C. zancles RHD as a promising candidate biocatalyst for marine PAH bioremediation.