Structural and Kinetic Properties of Liver Rhodanese from Coptodon zillii: Implications for Cyanide Detoxification in Gold Mining-Impacted Aquatic Ecosystems

The global gold extraction industry has been reported to use cyanide-based recovery processes, which pose environmental effects on water resources. The study examined <i>Coptodon zillii</i> liver rhodanese from a gold mining-impacted reservoir with a specific focus on the enzyme's critical function in cyanide detoxification. Rhodanese was purified using successive chromatographic techniques with 5.4 U/mg specific activity and 3.1-fold purification. The molecular weight of the native enzyme was 36 kDa, and the subunits were 17 kDa, indicative of a dimeric structure. Optimal enzymatic activity was recorded at pH 8.0 and 50 °C. The effect of metal ions was significantly varied: the activity was inhibited by BaCl₂, CaCl₂, NaCl, and MgCl₂, and KCl enhanced performance. The kinetic determinations showed Michaelis-Menten kinetics with a Km of 20.0 mM for sodium thiosulfate and 25.0 mM for potassium cyanide. The enzyme's minimal activity was identified toward 2-mercaptoethanol, ammonium persulfate, and ammonium sulfate, but with evidence of preference for thiosulfate utilization under the substrate specificity tests. The major interactions between the enzyme and the substrate were revealed by the molecular docking experiments. These showed Glu159, Gln161, and Arg173 formed important hydrogen bonds with thiosulfate, while Arg156 and Val172 were also involved. Other substrates are bound to Gln121 and Trp139 residues with much lower binding energy than thiosulfate. The findings increase our understanding of biochemical adaptation process knowledge in anthropogenically stressed environments, showing strategies of ecological resilience. The characterized enzymatic features showed potent cyanide detoxification potential, and the possible applications are in bioremediation strategies for mining-impacted aquatic ecosystems.