Premium
In vitro selection of optimal AbrB‐binding sites: comparison to known in vivo sites indicates flexibility in AbrB binding and recognition of three‐dimensional DNA structures
Author(s) -
Xu Ke,
Strauch Mark A.
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.1365-2958.1996.358882.x
Subject(s) - biology , flexibility (engineering) , dna , selection (genetic algorithm) , computational biology , in vivo , binding site , dna binding site , in vitro , genetics , gene , artificial intelligence , statistics , mathematics , gene expression , computer science , promoter
Summary The AbrB protein of Bacillus subtilis regulates expression of numerous genes, primarily through specific binding interactions to DNA regions containing transcriptional promoters. Although over 15 target regions for AbrB binding to chromosomally located sequences have been analysed by DNase I footprinting, no obvious consensus sequence or motif has yet emerged from their examination. Using in vitro selection techniques, we have isolated optimal AbrB‐binding sites from oligonucleotides containing 22 or 44 random base pairs. The best of these sites have an apparent in vitro K d which is fivefold lower than a similar‐sized DNA fragment containing the sequence corresponding to the AbrB‐binding site on the spoOE gene. We tested one of the sites in vivo and found that it confers AbrB‐mediated control upon a promoter not normally regulated by AbrB. In each of four separate trials, the selected sites possess motifs that converge to a simple consensus. It is argued that the nature and spacing of these motifs produce a type of three‐dimensional DNA structure recognizable by AbrB, and that known in vivo sites, which lack these motifs, possess an approximation of the optimal structural determinant.