Curation Information

Publication: Targets for the MalI repressor at the divergent Escherichia coli K-12 malX-malI promoters.;Lloyd GS, Godfrey RE, Busby SJ;FEMS microbiology letters 2010 Apr; 305(1):28-34 [20141531]
TF: MalI [P18811, view regulon]
Reported TF sp.: Escherichia coli str. K-12 substr. MG1655
Reported site sp.: Escherichia coli str. K-12 substr. MG1655
Created by: Erill Lab
Curation notes: -

Experimental Process

MalI binding sites were predicted by analogy to LacI (16 bp palindromic elements) [PMID:2670898]. The whole segment corresponding to the malI-malX intergenic region was mutagenized randomly by PCR and cloned. Colonies showing de-repressed activity in beta-gal assays for malI or malX were sequenced and consistent key mutations revealed the two sites for MalI. Multiple sequence alignments of the region with other Gammaproteobacteria further confirmed the results.

Transcription Factor Binding Sites

Sites reported in the paper
Sites matched in the genome

GGTAAAACGTTTTATC
GATAAAACGTTTTATC

Gene Regulation

Regulated genes for each binding site are displayed below. Gene regulation diagrams show binding sites, positively-regulated genes, negatively-regulated genes, both positively and negatively regulated genes, genes with unspecified type of regulation. For each indvidual site, experimental techniques used to determine the site are also given.

Site sequence	Regulated genes	Gene diagram	Experimental techniques	TF function	TF type
GGTAAAACGTTTTATC	malI,		Experimental technique details Beta-gal reporter assay Beta-gal reporter assay Reporter assay using the beta-galactosidase (lacZ) gene. The lacZ gene is typically fused to the promoter of interest. Differential regulation of the promoter mediated by the TF is assessed by induction of the system and evaluation of lacZ expression. Bacteria expressing lacZ appear blue when grown on a X-gal medium. The assay is often performed using a plasmid borne construction on a lacZ(def) strain. - Experimental technique details Multiple sequence alignment (MSA) (ECO:0005556) Multiple sequence alignment (MSA) - ECO:0005556 MSA can be used to align promoter regions and highlight regions of conservation that may be indicative of transcription factor binding sites. - Experimental technique details Random mutagenesis (ECO:0005530) Random mutagenesis - ECO:0005530 Non-targeted mutagenesis. Typically accomplished in vivo by means of radiation or a DNA-damaging agent, or in vitro using degenerate PCR. Not frequently used in TF-binding site determination. It is sometimes used to investigate the effect of mutations in transcription factor binding domains, in conjunction with expression assays. - Experimental technique details Visual sequence inspection (nan) Visual sequence inspection - nan Visual inspection of promoter region to identify putative binding sites based on previous knowledge -	repressor	dimer
GATAAAACGTTTTATC	malY, malX,		Experimental technique details Beta-gal reporter assay Beta-gal reporter assay Reporter assay using the beta-galactosidase (lacZ) gene. The lacZ gene is typically fused to the promoter of interest. Differential regulation of the promoter mediated by the TF is assessed by induction of the system and evaluation of lacZ expression. Bacteria expressing lacZ appear blue when grown on a X-gal medium. The assay is often performed using a plasmid borne construction on a lacZ(def) strain. - Experimental technique details Multiple sequence alignment (MSA) (ECO:0005556) Multiple sequence alignment (MSA) - ECO:0005556 MSA can be used to align promoter regions and highlight regions of conservation that may be indicative of transcription factor binding sites. - Experimental technique details Random mutagenesis (ECO:0005530) Random mutagenesis - ECO:0005530 Non-targeted mutagenesis. Typically accomplished in vivo by means of radiation or a DNA-damaging agent, or in vitro using degenerate PCR. Not frequently used in TF-binding site determination. It is sometimes used to investigate the effect of mutations in transcription factor binding domains, in conjunction with expression assays. - Experimental technique details Visual sequence inspection (nan) Visual sequence inspection - nan Visual inspection of promoter region to identify putative binding sites based on previous knowledge -	repressor	dimer