Hr. Beller et Am. Spormann, ANAEROBIC ACTIVATION OF TOLUENE AND O-XYLENE BY ADDITION TO FUMARATE IN DENITRIFYING STRAIN-T, Journal of bacteriology, 179(3), 1997, pp. 670-676
Anaerobic assays conducted with strain T, a denitrifying bacterium cap
able of mineralizing toluene to carbon dioxide, demonstrated that tolu
ene-grown, permeabilized cells catalyzed the addition of toluene to fu
marate to form benzylsuccinate. This reaction was not dependent on the
presence of coenzyme A (CoA) or ATP, In the presence of CoA, formatio
n of E-phenylitaconate from benzylsuccinate was also observed. Kinetic
studies demonstrated that the specific rate of benzylsuccinate format
ion from toluene and fumarate in assays with permeabilized cells was >
30% of the specific rate of toluene consumption in whole-cell suspensi
ons with nitrate; this observation suggests that benzylsuccinate forma
tion may be the first reaction in anaerobic toluene degradation by str
ain T. Use of deuterium-labeled toluene and gas chromatography-mass sp
ectrometry indicated that the H atom abstracted from the toluene methy
l group during addition to fumarate was retained in the succinyl moiet
y of benzylsuccinate. In this study, no evidence was found to support
previously proposed reactions of toluene with acetyl-CoA or succinyl-C
oA. Toluene-grown, permeabilized cells of strain T also catalyzed the
addition of o-xylene to fumarate to form (2-methylbenzyl) succinate. o
-Xylene is not a growth substrate for strain T, and its transformation
was probably cometabolic. With the exception of specific reaction rat
es, the observed characteristics of the toluene-fumarate addition reac
tion (i.e., retention of a methyl H atom and independence from CoA and
ATP) also apply to the o-xylene-fumarate addition reaction, Thus, add
ition to fumarate may be a biochemical strategy to anaerobically activ
ate a range of methylbenzenes.