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Effects of systematic variation of the minimal Escherichia coli met consensus operator site: in vivo and in vitro met repressor binding
Author(s) -
Wild Catherine M.,
McNally Teresa,
Phillips Simon E. V.,
Stockley Peter G.
Publication year - 1996
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1046/j.1365-2958.1996.00055.x
Subject(s) - biology , repressor , operator (biology) , in vitro , escherichia coli , in vivo , variation (astronomy) , genetics , computational biology , gene , transcription factor , physics , astrophysics
Summary We have produced a set of sequence variants based upon the idealized, minimal Escherichia coli met operator in which each position within the basic recognition unit, the 8bp met box (dAGACGTCT), has been changed to all other possible sequences containing single symmetrical base substitutions. The effects of these sequence variations have been assayed in vivo by monitoring the production of β‐galactosidase from a standard promoter regulated by the operator variants, and in vitro by gel‐retardation assay. The two sets of data are consistent and correlate well with expectations based on the three‐dimensional structure of the holorepressor bound to a minimal idealized operator and the results of in vitro evolution experiments. Comparison with two natural operators, metA and metC , suggests that in vivo , with non‐consensus operators, the repressor binds to at least four consecutive met boxes.