title: A transcription terminator in the groEx gene of symbiotic X-bacteria expressed in Escherichia coli.
abstract: The over-expressing groEx gene of symbiotic X-bacteria in Amoeba proteus has unique nucleotide motifs (Tx), containing two hairpins and a C-rich region at its 3'-end. To investigate the role of Tx as a transcription terminator, we mutated Tx and analyzed the effects on the expression of an upstream-located lacZ in E. coli. The level of beta-galactosidase (beta-gal) production in Tx deletion mutants was reduced to 23% of the control level. Site-directed mutation of the hairpin-1, C-rich region, and hairpin-2 reduced the beta-gal production to 28-64%, 33, and 20% of wild-type Tx, respectively. The amount of lacZ transcripts that were detected in RNA blots was proportional to the level of beta-gal. The Tx sequence had 97% termination efficiency in vivo, and the mutations in Tx resulted in read-through transcripts with significantly shortened half-life. In rho- E. coli, the level of the beta-gal production by Tx was reduced to 15% of that in rho+ E. coli. These results suggest that Tx is a Rho-dependent transcription terminator. Also, hairpin-2 is the most effective component among the three motifs of Tx for proper termination of the transcription and stability of mRNAs.
authors.affiliation: School of Biological Sciences, Seoul National University, Korea.
authors.forename: Jung Eun
authors.initials: JE
authors.lastname: Lee
authors.forename: Sung Han
authors.initials: SH
authors.lastname: Choi
authors.forename: Tae In
authors.initials: TI
authors.lastname: Ahn
mesh_terms: D000656:Amoeba / Q000382:microbiology; D000818:Animals; D001419:Bacteria / Q000235:genetics*; D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005798:Genes, Bacterial; D007763:Lac Operon; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D020871:RNA Stability; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013559:Symbiosis; D014158:Transcription, Genetic; D001616:beta-Galactosidase / Q000096:biosynthesis / Q000235:genetics
chemical_list: D012329:RNA, Bacterial; D001616:beta-Galactosidase