Characterization of Hexachlorocyclohexane Isomer Dehydrochlorination by LinA1 and LinA2 Using Multi-element Compound-Specific Stable Isotope Analysis.

Dehydrochlorination is one of the main (thus far discovered) processes for aerobic microbial transformation of hexachlorocyclohexane (HCH) which is mainly catalyzed by LinA enzymes. In order to gain a better understanding of the reaction mechanisms, multi-element compound-specific stable isotope analysis was applied for evaluating α- and γ-HCH transformations catalyzed by LinA1 and LinA2 enzymes. The isotopic fractionation (ε) values for particular elements of (+)α-HCH (ε = -10.8 ± 1.0‰, ε = -4.2 ± 0.5‰, ε = -154 ± 16‰) were distinct from the values for (-)α-HCH (ε = -4.1 ± 0.7‰, ε = -1.6 ± 0.2‰, ε = -68 ± 10‰), whereas the dual-isotope fractionation patterns were almost identical for both enantiomers (Λ = 2.4 ± 0.4 and 2.5 ± 0.2, Λ = 12.9 ± 2.4 and 14.9 ± 1.1). The ε of γ-HCH transformation by LinA1 and LinA2 were -7.8 ± 1.0‰ and -7.5 ± 0.8‰ (ε), -2.7 ± 0.3‰ and -2.5 ± 0.4‰ (ε), -170 ± 25‰ and -150 ± 13‰ (ε), respectively. Similar Λ values (2.7 ± 0.2 and 2.9 ± 0.2) were observed as well as similar Λ values (20.1 ± 2.0 and 18.4 ± 1.9), indicating a similar reaction mechanism by both enzymes during γ-HCH transformation. This is the first data set on 3D isotope fractionation of α- and γ-HCH enzymatic dehydrochlorination, which gave a more precise characterization of the bond cleavages, highlighting the potential of multi-element compound-specific stable isotope analysis to characterize different transformation processes (e.g., dehydrochlorination and reductive dehalogenation).