ISSN:
1617-4623
Keywords:
Bacillus subtilis
;
gnt operon
;
Transcriptional repressor
;
Insertional operator mutation
;
GntR family
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
Notes:
Abstract TheBacillus subtilis gnt operon is negatively regulated via interaction of thegnt repressor (GntR) with an operator upstream ofgntR, which is antagonized by gluconate. An 8 bp insertional operator mutation (gntOi) of thegnt operon was constructed which affected the expression level of this operon. Two suppressors of thisgntOi mutation, exhibiting normal expression, were also isolated; one involved a threonine substitution for the Ala-48 residue (gntR48T) within the helix-turn-helix DNA-binding motif of GntR, and the other an adenine substitution for the guanine at nucleotide — 4 within thegntOi operator (gntOiM4A) (+1 is the transcription initiation site). ThegntR48T mutation by itself rendered thegnt operon partially constitutive. When thegntR43L mutation, which renders thegnt operon fully constitutive, was introduced into thegntOi orgntOiM4A mutant, the operator mutations were found not to affect the promoter activity of thegnt operon. These in vivo results indicate that thegntOi mutation affects the operator interaction with GntR, causing a low expression level even in the presence of gluconate. In vitro gel retardation and DNase I footprint analyses demonstrated that even when gluconate was present, GntR still bound to thegntOi operator region.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF02423454
Permalink
