title: Characterization of FadR, a global transcriptional regulator of fatty acid metabolism in Escherichia coli. Interaction with the fadB promoter is prevented by long chain fatty acyl coenzyme A.
abstract: The Escherichia coli fadR gene product, FadR, is a multifunctional regulator of fatty acid metabolism. In this work we have purified FadR by a two-step procedure employing two ion-exchange columns. The amino-terminal sequence of the purified protein confirms the sequence of the protein derived from analysis of the DNA sequence (DiRusso, C. C. (1988) Nucleic Acids Res. 16, 7995-8009) and indicates that the initiating methionine is cleaved from the mature protein. Purified FadR binds to a 326-base pair HaeIII fragment of fadB DNA which carries the fadB promoter. DNase I footprinting localizes the operator to a sequence, 5' ATCTGGTACGACCAGAT 3', at +1 to +17 nucleotides relative to the start of transcription. Using protein-DNA gel retention assays, we estimate the Keq of FadR binding to the fadB operator to be approximately 3 x 10(-10) M. Binding of FadR is specifically inhibited by long chain fatty acyl-CoA compounds. The apparent Ki values for oleoyl-CoA, palmitoyl-CoA, and palmitoleoyl-CoA are each 5 nM while that of myristoyl-CoA is 250 nM. Decanoyl-CoA, crotonoyl-CoA, and free fatty acids inhibited binding only at concentrations above 1 microM.
authors.affiliation: Department of Biochemistry, University of Tennessee, Memphis 38163.
authors.forename: C C
authors.initials: CC
authors.lastname: DiRusso
authors.forename: T L
authors.initials: TL
authors.lastname: Heimert
authors.forename: A K
authors.initials: AK
authors.lastname: Metzger
mesh_terms: D000214:Acyl Coenzyme A / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D002853:Chromatography, Liquid; D016172:DNA Fingerprinting; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005227:Fatty Acids / Q000378:metabolism*; D008969:Molecular Sequence Data; D009875:Operator Regions, Genetic; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins; D014158:Transcription, Genetic
chemical_list: D000214:Acyl Coenzyme A; D001426:Bacterial Proteins; D004269:DNA, Bacterial; C057119:FadR protein, Bacteria; D005227:Fatty Acids; D012097:Repressor Proteins