Isotope Fractionation Pinpoints Membrane Permeability as a Barrier to Atrazine Biodegradation in Gram-negative Polaromonas sp. Nea-C

Biodegradation of persistent pesticides like atrazine often stalls at low concentrations in the environment. While mass transfer does not limit atrazine degradation by the Gram-positive Arthrobacter aurescens TC1 at high concentrations (>1 mg/L), evidence of bioavailability limitations is emerging at trace concentrations (<0.1 mg/L). To assess the bioavailability constraints on biodegradation, the roles of cell wall physiology and transporters remain imperfectly understood. Here, compound-specific isotope analysis (CSIA) demonstrates that cell wall physiology (i.e., the difference between Gram-negative and Gram-positive bacteria) imposes mass transfer limitations in atrazine biodegradation even at high concentrations. Atrazine biodegradation by Gram-negative Polaromonas sp. Nea-C caused significantly less isotope fractionation (ϵ(C) = -3.5 ‰) than expected for hydrolysis by the enzyme TrzN (ϵ(C) = -5.0 ‰) and observed in Gram-positive Arthrobacter aurescens TC1 (ϵ(C) = -5.4 ‰). Isotope fractionation was recovered in cell-free extracts (ϵ(C) = -5.3 ‰) where no cell envelope restricted pollutant uptake. When active transport was inhibited with cyanide, atrazine degradation rates remained constant demonstrating that atrazine mass transfer across the cell envelope does not depend on active transport but is a consequence of passive cell wall permeation. Taken together, our results identify the cell envelope of the Gram-negative bacterium Polaromonas sp. Nea-C as a relevant barrier for atrazine biodegradation.