title: Analysis of the tryptophanase expression in Symbiobacterium thermophilum in a coculture with Geobacillus stearothermophilus.
abstract: The tryptophanase-positive Symbiobacterium thermophilum is a free-living syntrophic bacterium that grows effectively in a coculture with Geobacillus stearothermophilus. Our studies have shown that S. thermophilum growth depends on the high CO2 and low O2 condition established by the precedent growth of G. stearothermophilus. The use of an anoxic atmosphere containing high CO2 allows S. thermophilum to grow independently of G. stearothermophilus, but the cellular yield is ten times lower than that achieved in the coculture. In this study, we characterized the coculture-dependent expression and activity of tryptophanase in S. thermophilum. S. thermophilum cells accumulated a marked amount of indole in a coculture with G. stearothermophilus, but not in the bacterium's pure culture irrespective of the addition of tryptophan. S. thermophilum cells accumulated indole in its pure culture consisting of conditioned medium (medium supplied with culture supernatant of G. stearothermophilus). Proteomic analysis identified the protein specifically produced in the S. thermophilum cells grown in conditioned medium, which was a tryptophanase encoded by tna2 (STH439). An attempt to isolate the tryptophanase-inducing component from the culture supernatant of G. stearothermophilus was unsuccessful, but we did discover that the indole accumulation occurs when 10 mM bicarbonate is added to the medium. RT-PCR analysis showed that the addition of bicarbonate stimulated transcription of tna2. The transcriptional start site, identified within the tna2 promoter, was preceded by the -24 and -12 consensus sequences specified by an alternative sigma factor, σ(54). The evidence suggests that the transcription of some genes involved in amino acid metabolism is σ(54)-dependent, and that a bacterial enhancer-binding protein containing a PAS domain controls the transcription under the presence of high levels of bicarbonate.
authors.affiliation: Institute of Biogeosciences, Japan Agency for Marine-Earth Science and Technology (JAMSTEC), 2-15 Natsushima-cho, Yokosuka, Kanagawa, 237-0061, Japan.
authors.forename: Tomo-O
authors.initials: TO
authors.lastname: Watsuji
authors.forename: Hideaki
authors.initials: H
authors.lastname: Takano
authors.forename: Tomoya
authors.initials: T
authors.lastname: Yamabe
authors.forename: Satoshi
authors.initials: S
authors.lastname: Tamazawa
authors.forename: Hiroka
authors.initials: H
authors.lastname: Ikemura
authors.forename: Takanori
authors.initials: T
authors.lastname: Ohishi
authors.forename: Tohyo
authors.initials: T
authors.lastname: Matsuda
authors.forename: Hatsumi
authors.initials: H
authors.lastname: Shiratori-Takano
authors.forename: Teruhiko
authors.initials: T
authors.lastname: Beppu
authors.forename: Kenji
authors.initials: K
authors.lastname: Ueda
mesh_terms: D001665:Binding Sites; D002245:Carbon Dioxide / Q000378:metabolism; D003470:Culture Media / Q000737:chemistry; D020869:Gene Expression Profiling; D006094:Gram-Positive Bacteria / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development*; D007211:Indoles / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D020543:Proteome / Q000032:analysis; D051877:RNA Polymerase Sigma 54 / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D024363:Transcription Initiation Site; D014368:Tryptophanase / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D003470:Culture Media; D007211:Indoles; D020543:Proteome; D002245:Carbon Dioxide; C030374:indole; D051877:RNA Polymerase Sigma 54; D014368:Tryptophanase; D010100:Oxygen