Abstract
The metJ gene encoding the methionine aporepressor was placed under the control of a strong and inducible promoter, ptac. Bacterial strains carrying the recombinant plasmid pIP35 overproduced the regulatory protein by a factor of 200 over the wild type strain as determined by the immunoblot technique. The purified metJ gene product negatively controls the expression of the metF gene, in a cell-free system as shown by repression of beta-galactosidase synthesis under the control of the metF promoter. The metJ protein binds to a DNA fragment containing the potential operator of the metF gene with an affinity which is 10 times greater in the presence of S-adenosylmethionine than in its absence. Equilibrium dialysis experiments showed that the met aporepressor binds 2 mol of S-adenosylmethionine per mol of dimer with a dissociation constant of 200 microM.
Highlights
The methionine aporepressor, the polypeptide product of t h e metJ gene, controls seven unlinked transcriptional units in Escherichia coli and Salmonella typhimurium: metA, metBL, metC, me@, metF, the biosynthetic genes €or methionine; metK, responsible for the biosynthesis of S-adenosylmethionine; and metJ, the structural gene forthe m e t aporepressor [1,2,3,4,5]
When themetJ protein overproducing strain the product of the metJ gene we have purified behaves like (GT809) was grown in minimal medium in the presence of IPTG, 3 timesmore protein was detected than in the absence an efficient aporepressor of the metFtranscriptional unit with S-adenosylmethionine as a corepressor, in agreement with of IPTG
We metF Gene-To test the biological activity of the purified think that repression at a relatively high concentration of metJ protein, we used a Zubay typein vitro systempro- methionine is due to theformation of S-adenosylmethionine grammed with DNA from plasmid pIP36
Summary
The methionine aporepressor, the polypeptide product of t h e metJ gene, controls seven unlinked transcriptional units in Escherichia coli and Salmonella typhimurium: metA, metBL, metC, me@, metF, the biosynthetic genes €or methionine; metK, responsible for the biosynthesis of S-adenosylmethionine; and metJ, the structural gene forthe m e t aporepressor [1,2,3,4,5]. No radiochemically labeled pure proteinwas needed as a tracer,the met aporepressor being already detectable on a SDS polyacrylamide gel in crude extracts of the highly overproducing strain GT809.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
