Competitive inhibition underlies Cr(III) toxicity to soil arylsulfatase: Kinetic transitions during aging and molecular validation of detoxification mechanisms.

Soil arylsulfatase (ARS) plays a critical role in sulfur cycling by catalyzing the hydrolysis of organic sulfates and serves as a sensitive bioindicator of heavy metal toxicity, including chromium (Cr(III)). However, its response to Cr(III) stress remains largely unexplored. This study integrates ecotoxicological and molecular approaches to explore interactions among soil properties, recent/aged Cr(III) contamination, and ARS kinetics across five soil types. Results show that soil organic matter and total nitrogen influence enzyme distribution: extracellular dominance in red, paddy, and saline soils versus intracellular prevalence in black and alluvial soils. Recent Cr(III) exposure induces competitive inhibition (≤70% activity loss; Kic = 0.20-2.32 mM), reducing substrate affinity, with the most severe effects in the total and extracellular fractions. Intracellular ARS emerged as the most sensitive kinetic biomarker, exhibiting significantly lower ecological dose values (ED10, p < 0.05) and up to 37-fold higher inhibition variability than other fractions. Aging (0-90 days) induced kinetic shifts: SOM-rich black soil transitioned to non-competitive inhibition, while alluvial soil exhibited mixed-type inhibition. The dominance of competitive inhibition (Km > Kic > Kiu) suggests stable enzyme-inhibitor complex formation. Molecular docking confirmed Cr(III) binding at the catalytic site (-1.862 kcal mol-1), supported by FTIR spectra showing an N-H blueshift (3170 → 3380 cm-1) indicating coordination with N-H groups. Humic acid reduced Cr(III) toxicity via steric hindrance (-6.800 kcal mol-1), lowering Cr(III) binding affinity by 24% (-2.312 kcal mol-1). These findings reveal mechanistic insights into Cr(III) inhibition and organic-matter-mediated mitigation, supporting more precise ecological risk assessments.