Combined C and Cl Isotope Effects Indicate Differences between Corrinoids and Enzyme (<i>Sulfurospirillum multivorans</i> PceA) in Reductive Dehalogenation of Tetrachloroethene, But Not Trichloroethene

Abstract

The role of the corrinoid cofactor in reductive dehalogenation catalysis by tetrachloroethene reductive dehalogenase (PceA) of <i>Sulfurospirillum multivorans</i> was investigated using isotope analysis of carbon and chlorine. Crude extracts containing PceAharboring either a native <i>norpseudo</i>-B₁₂ or the alternative <i>nor</i>-B₁₂ cofactorwere applied for dehalogenation of tetrachloroethene (PCE) or trichloroethene (TCE), and compared to abiotic dehalogenation with the respective purified corrinoids (<i>norpseudo</i>vitamin B₁₂ and <i>nor</i>vitamin B₁₂), as well as several commercially available cobalamins and cobinamide. Dehalogenation of TCE resulted in a similar extent of C and Cl isotope fractionation, and in similar dual-element isotope slopes (ε_C/ε_Cl) of 5.0–5.3 for PceA enzyme and 3.7–4.5 for the corrinoids. Both observations support an identical reaction mechanism. For PCE, in contrast, observed C and Cl isotope fractionation was smaller in enzymatic dehalogenation, and dual-element isotope slopes (2.2–2.8) were distinctly different compared to dehalogenation mediated by corrinoids (4.6−7.0). Remarkably, ε_C/ε_Cl of PCE depended in addition on the corrinoid type: ε_C/ε_Cl values of 4.6 and 5.0 for vitamin B₁₂ and <i>nor</i>vitamin B₁₂ were significantly different compared to values of 6.9 and 7.0 for <i>norpseudo</i>vitamin B₁₂ and dicyanocobinamide. Our results therefore suggest mechanistic and/or kinetic differences in catalytic PCE dehalogenation by enzymes and different corrinoids, whereas such differences were not observed for TCE