Cloning, nucleotide sequence, and overexpression in Escherichia coli of the ?-tyrosinase gene from an obligately symbiotic thermophile, Symbiobacterium thermophilum

Abstract

Symbiobacterium thermophilum is an obligately symbiotic thermophile that can grow only in coculture with a specific Bacillus strain. The amino acid sequences of fragments obtained by cyanogen bromide decomposition of the thermostable beta-tyrosinase (tyrosine phenol-lyase, E.C. 4.1.99.2) from this organism resembled that of the tryptophanase produced by the same organism. DNA-probing with the tryptophanase gene as the hybridization probe led to cloning in Escherichia coli of the beta-tyrosinase (tpl) gene. The nucleotide sequence revealed that the beta-tyrosinase of 458 amino acids (relative molecular mass, 52269) showed significant similarity in amino acid sequence to the tryptophanase over the entire sequence. DNA manipulation of the cloned tpl gene in E. coli led to production of 375 times as much beta-tyrosinase as that produced by the original S. thermophilum strain.

Lessons from Studies ofSymbiobacteriumthermophilum, a Unique Syntrophic Bacterium

“…Gramstaining of the cell was negative. The production of tryptophanase 3) and -tyrosinase 4) is one of the characteristics of enteric bacteria. In addition, the cell surface structure of S. thermophilum consisted of multiple layers distinct from that of Gram-positive bacteria.…”

Comparative genomic analysis by microbial COGs self-attraction rate

“…(Ueda et al, 2004). The ability of S.thermophilum leading to tryptophanase and tyrosinase production, typical of enterobacteria, further suggests that this bacterium may be considered a Gram negative (Hirahara et al, 1992;Hirahara et al, 1993).…”

Symbiobacterium thermophilum gen. nov., sp. nov., a symbiotic thermophile that depends on co-culture with a Bacillus strain for growth.

“…Keywords : Symbiobacterium thermophilum, 16S rDNA, tryptophanase, high-GjC Gram-positive bacteria A thermophilic bacterium (strain T T ) producing tryptophanase and tyrosine-phenyl lyase ( β-tyrosinase) was obtained from compost at 60 mC in mixed culture with a thermophilic Bacillus strain (Suzuki et al, 1988) and was used as a source of these distinctly heat-stable enzymes (Suzuki et al, 1991Hirahara et al, 1992Hirahara et al, , 1993. We originally isolated these organisms from a sample collected in Japan ; isolation of similar organisms in Korea was also reported by Lee et al (1997).…”