Abstract
Certain mutations in gyrA and gyrB, the genes encoding the two subunits of DNA gyrase, are known to influence expression of the his operon (K. E. Rudd and R. Menzel, Proc. Natl. Acad. Sci. USA 84:517-521, 1987). Such mutations lead to a decrease in tRNA(His) levels and consequently to an attenuator- dependent increase in his operon expression. This effect presumably is due to the dependence of the hisR promoter (hisR encodes tRNA(His)) on supercoiling for maximal activity. We used a relaxed (Rel-) strain of Escherichia coli to isolate gyrB mutants by selecting for resistance to the histidine antimetabolite 3-amino-1,2,4-triazole and then screening for temperature- sensitive growth on rich medium. Rel- mutants, which generally have lower basal levels of ppGpp (a positive regulator of his operon transcription), are more sensitive than wild-type E. coli to aminotriazole. The chance of isolating spoT mutants, which can be selected with a similar procedure, was decreased by selecting in the presence of a multicopy plasmid that carries the wild-type spoT gene. Under these conditions, gyrB mutants were isolated preferentially. This scheme selects for loss of function of DNA gyrase, rather than for its alteration due to resistance to specific gyrase inhibitors, and thus a greater variety of gyrase mutations might be obtainable.
| Original language | English |
|---|---|
| Pages (from-to) | 5479-5481 |
| Number of pages | 3 |
| Journal | Journal of Bacteriology |
| Volume | 174 |
| Issue number | 16 |
| State | Published - Jan 1 1992 |
| Externally published | Yes |
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ASJC Scopus subject areas
- Applied Microbiology and Biotechnology
- Immunology
Cite this
Mutations causing aminotriazole resistance and temperature sensitivity reside in gyrB, which encodes the B subunit of DNA gyrase. / Toone, W. M.; Rudd, K. E.; Friesen, J. D.
In: Journal of Bacteriology, Vol. 174, No. 16, 01.01.1992, p. 5479-5481.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Mutations causing aminotriazole resistance and temperature sensitivity reside in gyrB, which encodes the B subunit of DNA gyrase
AU - Toone, W. M.
AU - Rudd, K. E.
AU - Friesen, J. D.
PY - 1992/1/1
Y1 - 1992/1/1
N2 - Certain mutations in gyrA and gyrB, the genes encoding the two subunits of DNA gyrase, are known to influence expression of the his operon (K. E. Rudd and R. Menzel, Proc. Natl. Acad. Sci. USA 84:517-521, 1987). Such mutations lead to a decrease in tRNA(His) levels and consequently to an attenuator- dependent increase in his operon expression. This effect presumably is due to the dependence of the hisR promoter (hisR encodes tRNA(His)) on supercoiling for maximal activity. We used a relaxed (Rel-) strain of Escherichia coli to isolate gyrB mutants by selecting for resistance to the histidine antimetabolite 3-amino-1,2,4-triazole and then screening for temperature- sensitive growth on rich medium. Rel- mutants, which generally have lower basal levels of ppGpp (a positive regulator of his operon transcription), are more sensitive than wild-type E. coli to aminotriazole. The chance of isolating spoT mutants, which can be selected with a similar procedure, was decreased by selecting in the presence of a multicopy plasmid that carries the wild-type spoT gene. Under these conditions, gyrB mutants were isolated preferentially. This scheme selects for loss of function of DNA gyrase, rather than for its alteration due to resistance to specific gyrase inhibitors, and thus a greater variety of gyrase mutations might be obtainable.
AB - Certain mutations in gyrA and gyrB, the genes encoding the two subunits of DNA gyrase, are known to influence expression of the his operon (K. E. Rudd and R. Menzel, Proc. Natl. Acad. Sci. USA 84:517-521, 1987). Such mutations lead to a decrease in tRNA(His) levels and consequently to an attenuator- dependent increase in his operon expression. This effect presumably is due to the dependence of the hisR promoter (hisR encodes tRNA(His)) on supercoiling for maximal activity. We used a relaxed (Rel-) strain of Escherichia coli to isolate gyrB mutants by selecting for resistance to the histidine antimetabolite 3-amino-1,2,4-triazole and then screening for temperature- sensitive growth on rich medium. Rel- mutants, which generally have lower basal levels of ppGpp (a positive regulator of his operon transcription), are more sensitive than wild-type E. coli to aminotriazole. The chance of isolating spoT mutants, which can be selected with a similar procedure, was decreased by selecting in the presence of a multicopy plasmid that carries the wild-type spoT gene. Under these conditions, gyrB mutants were isolated preferentially. This scheme selects for loss of function of DNA gyrase, rather than for its alteration due to resistance to specific gyrase inhibitors, and thus a greater variety of gyrase mutations might be obtainable.
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UR - http://www.scopus.com/inward/citedby.url?scp=0026760614&partnerID=8YFLogxK
M3 - Article
C2 - 1322887
AN - SCOPUS:0026760614
VL - 174
SP - 5479
EP - 5481
JO - Journal of Bacteriology
JF - Journal of Bacteriology
SN - 0021-9193
IS - 16
ER -