1. Polyribosomes and RNA were isolated from cultures in which tryptophanase (EC 4.2.1.–) was induced. The polyribosomes were incubated under conditions of protein synthesis, in the presence of a radioactive amino acid and a post-ribosomal supernatant fraction obtained from repressed cells. The RNA preparations were incubated under conditions of protein synthesis in the presence of a radioactive amino acid and a supernatant fraction containing ribosomes from repressed cells. 2. The system was characterized and the synthesis of a radioactive protein with the same chromatographic properties as tryptophanase was demonstrated. This synthesis was shown to be time-dependent and required the presence of RNA from induced cultures, ribosomes and an energy supply; it was inhibited by chloramphenicol. 3. The maximum activity for the synthesis of this protein was found to be associated with 23S rRNA isolated from sucrose gradients. 4. The N-terminal amino acid of tryptophanase was labelled in the protein synthesized in this system but not in the protein synthesized by polyribosomes (without added RNA). Conversely, the C-terminal amino acid of tryptophanase was labelled in the polyribosome system but not in the RNA-containing system. 5. Tryptic digests of protein labelled in vitro were compared with those of tryptophanase. No labelled tryptic peptides were identified other than tryptophanase tryptic peptides. An analysis of the results implied that in the polyribosome system almost the complete tryptophanase subunit chain was labelled but that in the RNA-containing system these chains were incompletely synthesized. 6. Sucrose-gradient analysis of protein synthesized in the RNA-containing system suggested that it cannot be converted into structures with the same sedimentation properties as native tryptophanase. 7. The significance of these results for the assay of tryptophanase mRNA and for an understanding of the control of the translation of this mRNA in vivo is discussed.
Skip Nav Destination
Article navigation
November 1971
-
Cover Image
Cover Image
- PDF Icon PDF LinkFront Matter
- PDF Icon PDF LinkTable of Contents
- PDF Icon PDF LinkAdvertising
Research Article|
November 01 1971
Cell-free synthesis of tryptophanase from Escherichia coli. Use of ribonucleic acid isolated from induced cells and a comparison of the product from a system employing ribosomes with that from one employing ribosomes and exogenous ribonucleic acid
J. H. Parish;
J. H. Parish
1Department of Biochemistry, University of Leeds, 9 Hyde Terrace, Leeds LS2 9LS, U.K.
Search for other works by this author on:
S. A. M. Khairul Bashar;
S. A. M. Khairul Bashar
1Department of Biochemistry, University of Leeds, 9 Hyde Terrace, Leeds LS2 9LS, U.K.
Search for other works by this author on:
N. L. Brown;
N. L. Brown
1Department of Biochemistry, University of Leeds, 9 Hyde Terrace, Leeds LS2 9LS, U.K.
Search for other works by this author on:
Marjorie Brown
Marjorie Brown
1Department of Biochemistry, University of Leeds, 9 Hyde Terrace, Leeds LS2 9LS, U.K.
Search for other works by this author on:
Publisher: Portland Press Ltd
© 1971 The Biochemical Society
1971
Biochem J (1971) 125 (2): 643–653.
Citation
J. H. Parish, S. A. M. Khairul Bashar, N. L. Brown, Marjorie Brown; Cell-free synthesis of tryptophanase from Escherichia coli. Use of ribonucleic acid isolated from induced cells and a comparison of the product from a system employing ribosomes with that from one employing ribosomes and exogenous ribonucleic acid. Biochem J 1 November 1971; 125 (2): 643–653. doi: https://doi.org/10.1042/bj1250643
Download citation file:
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Could not validate captcha. Please try again.