Identification and enzymic analysis of a novel protein associated with production of hydrogen sulfide and L-serine from L-cysteine in Fusobacterium nucleatum subsp. nucleatum ATCC 25586

Abstract

A third enzyme that produces hydrogen sulfide from L-cysteine was identified in Fusobacterium nucleatum subsp. nucleatum. The fn1055 gene was cloned from a cosmid library constructed with genomic DNA of F. nucleatum ATCC 25586. Despite the database annotation that the product of fn1055 is a cysteine synthase, reverse-phase HPLC revealed that no L-cysteine was produced in vitro by the purified Fn1055 protein; however, the enzyme did produce L-serine. In addition, a cysteine auxotroph, Escherichia coli NK3, transformed with a plasmid containing the fn1055 gene did not grow without cysteine, which further suggests that Fn1055 does not function as a cysteine synthase. The Michaelis-Menten kinetics (K(m) =0.09 ± 0.001 mM and k(cat) =5.43 ± 0.64 s(-1)) of the purified enzyme showed that the capacity of Fn1055 to produce hydrogen sulfide was between that of two other enzymes, Fn0625 and Fn1220. Incubation of Fn1055 with L-cysteine resulted in the production of hydrogen sulfide, but not of pyruvate, ammonia or lanthionine, which are all byproducts produced in addition to hydrogen sulfide when Fn0625 or Fn1220 is incubated with L-cysteine. Instead, Fn1055 produced L-serine in its reaction with L-cysteine. Fn1055 produces hydrogen sulfide from l-cysteine by a mechanism that is different from that of Fn0625 or Fn1220.