Regulation of the Escherichia coli tna operon: nascent leader peptide control at the tnaC stop codon.
Expression of the tryptophanase (tna) operon of Escherichia coli is regulated by catabolite repression and by tryptophan-induced transcription antitermination at Rho-dependent termination sites in the leader region of the operon. Tryptophan induction is dependent on translation of a short leader peptide coding region, tnaC, that contains a single, crucial tryptophan codon. Recent studies suggest that during induction, the TnaC leader peptide acts in cis on the translating ribosome to inhibit its release at the tnaC stop codon. In the present study we use a tnaC-UGA-'lacZ construct lacking the tnaC-tnaA spacer region to analyze the effect of TnaC synthesis on the behavior of the ribosome that translates tnaC. The tnaC-UGA-'lacZ construct is not expressed significantly in the presence or absence of inducer. However, it is expressed in the presence of UGA suppressors, or when the structural gene for polypeptide release factor 3 is disrupted, or when wild-type tRNATrP is overproduced. In each situation, tnaC-UGA-'lacZ expression is reduced appreciably by the presence of inducing levels of tryptophan. Replacing the tnaC UGA stop codon with a sense codon allows considerable expression, which is also reduced, although to a lesser extent, by the addition of tryptophan. Inhibition by tryptophan is not observed when Trp codon 12 of tnaC is changed to a Leu codon. Overexpression of tnaC in trans from a multicopy plasmid prevents inhibition of expression by tryptophan. These results support the hypothesis that the TnaC leader peptide acts in cis to alter the behavior of the translating ribosome.
Konan
K V
KV
Department of Biological Sciences, Stanford University, California 94305-5020, USA.
Yanofsky
C
C
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D001483:Base Sequence; D018388:Codon, Terminator; D003470:Culture Media; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009876:Operon; D010454:Peptide Termination Factors / Q000235:genetics / Q000502:physiology; D021382:Protein Sorting Signals / Q000502:physiology; D011994:Recombinant Proteins; D012234:Rho Factor / Q000235:genetics / Q000502:physiology; D013489:Suppression, Genetic; D014364:Tryptophan / Q000378:metabolism; D014368:Tryptophanase / Q000235:genetics*
D001426:Bacterial Proteins; D018388:Codon, Terminator; D003470:Culture Media; D029968:Escherichia coli Proteins; D010454:Peptide Termination Factors; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D012234:Rho Factor; C087971:peptide-chain-release factor 3; C081473:tnaC protein, E coli; D014364:Tryptophan; D014368:Tryptophanase