title: Isolation and characterization of a moderate thermophile, Mycobacterium phlei GTIS10, capable of dibenzothiophene desulfurization.
abstract: An organism, identified as Mycobacterium phlei GTIS10, was isolated based on its ability to use dibenzothiophene (DBT) as a sole source of sulfur for growth at 30-52 degrees C. Similar to other biodesulfurization-competent organisms, M. phlei GTIS10 converts DBT to 2-hydroxybiphenyl (2-HBP), as detected by HPLC. The specific desulfurization activity of the 50 degrees C M. phlei GTIS10 culture was determined to be 1.1+/-0.07 micromol 2-HBP min(-1) (g dry cell)(-1). M. phlei GTIS10 can also utilize benzothiophene and thiophene as sulfur sources for growth. The dszABC operon of M. phlei GTIS10 was cloned and sequenced and was found to be identical to that of Rhodococcus erythropolis IGTS8. The presence of the R. erythropolis IGTS8 120-kb plasmid pSOX, which encodes the dszABC operon, has been demonstrated in M. phlei GTIS10. Even though identical dsz genes are contained in both cultures, the temperature at which resting cells of R. erythropolisIGTS8 reach the highest rate of DBT metabolism is near 30 degrees C whereas the temperature that shows the highest activity in resting cell cultures of M. phlei GTIS10 is near 50 degrees C, and activity is detectable at temperatures as high as 57 degrees C. In M. phlei GTIS10, the rate-limiting step in vivo appears to be the conversion of DBT to dibenzothiophene sulfone catalyzed by the product of the dszC gene, DBT monooxygenase. The thermostability of individual desulfurization enzymes was determined and 2-hydroxybiphenyl-2-sulfinate sulfinolyase, encoded by dszB, was found to be the most thermolabile. These results demonstrate that the thermostability of individual enzymes determined in vitro is not necessarily a good predictor of the functional temperature range of enzymes in vivo.
authors.affiliation: Gas Technology Institute, 1700 South Mount Prospect Road, Des Plaines, IL 60018, USA.
authors.forename: K J
authors.initials: KJ
authors.lastname: Kayser
authors.forename: L
authors.initials: L
authors.lastname: Cleveland
authors.forename: H-S
authors.initials: HS
authors.lastname: Park
authors.forename: J-H
authors.initials: JH
authors.lastname: Kwak
authors.forename: A
authors.initials: A
authors.lastname: Kolhatkar
authors.forename: J J
authors.initials: JJ
authors.lastname: Kilbane
mesh_terms: D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics*; D006358:Hot Temperature; D009161:Mycobacterium / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics; D010105:Oxygenases / Q000737:chemistry / Q000235:genetics; D016133:Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D013876:Thiophenes / Q000378:metabolism*
chemical_list: D004269:DNA, Bacterial; D013876:Thiophenes; D010088:Oxidoreductases; D010105:Oxygenases; C421210:dibenzothiophene sulfone monooxygenase; C090100:dibenzothiophene monooxygenase; C016366:dibenzothiophene