Regulation of the nap operon encoding the periplasmic nitrate reductase of Paracoccus pantotrophus: delineation of DNA sequences required for redox control.
Expression of the nap operon, encoding the periplasmic nitrate reductase in Paracoccus pantotrophus, is maximal when cells are grown aerobically, but not anaerobically, with butyrate. Two promoters, termed P1 and P2, control operon expression and the operon-proximal P2 promoter is primarily responsible for increased nap expression in the presence of butyrate. A near-perfect palindromic sequence is centred at +7, relative to the P2 transcription start site. Mutation of this palindrome demonstrated that it is important for regulation of nap operon expression in response to both the redox and the oxidation state of the carbon substrate. A 5' deletion analysis of the nap promoter fused to lacZ revealed that full redox control of expression was retained when the DNA sequence up to position -49 bp, relative to the operon-distal P1 transcription start site, was removed. Encroaching beyond this position resulted in an approximately 4-fold reduction in expression when cells were grown aerobically with butyrate. Additionally, point mutations at position -38 and -45 relative to P1 also resulted in a reduction in expression during aerobic growth with butyrate. A GC-rich region of nap promoter DNA, centred on position -41 relative to the P1 transcription start site is thus proposed as a second DNA motif that is important for efficient expression of the nap operon.
Ellington
M J K
MJ
Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, UK.
Fosdike
W L J
WL
Sawers
R G
RG
Richardson
D J
DJ
Ferguson
S J
SJ
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002087:Butyrates / Q000378:metabolism; D003470:Culture Media; D015964:Gene Expression Regulation, Bacterial; D016297:Mutagenesis, Site-Directed; D050901:Nitrate Reductase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009876:Operon; D010084:Oxidation-Reduction; D042084:Paracoccus pantotrophus / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D019897:Periplasm / Q000201:enzymology*; D011401:Promoter Regions, Genetic; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D002087:Butyrates; D003470:Culture Media; D050901:Nitrate Reductase