Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.6 (RNA polymerase)
 34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Pseudomonas putida strain DS1 utilizes dimethyl sulfide (DMS) as a sulfur source, and desulfurizes it via dimethyl sulfoxide (DMSO), dimethyl sulfone (DMSO(2)) and methanesulfonate (MSA). Its Tn5 mutant, Dfi74J, no longer utilized DMS, DMSO and DMSO(2), but could oxidize DMS to DMSO(2), suggesting that the conversion of DMSO(2) to MSA was interrupted in the mutant. Sequencing of the Tn5 flanking region of Dfi74J demonstrated that a gene, sfnR (designated for dimethyl sulfone utilization), encoding a transcriptional regulator containing an ATP-dependent sigma(54)-association domain and a DNA-binding domain, was disrupted. sfnR is part of an operon with two other genes, sfnE and sfnC, located immediately upstream of sfnR and in the same orientation. The genes encode NADH-dependent FMN reductase (SfnE) and FMNH(2)-dependent monooxygenase (SfnC). Complementation of Dfi74J with an sfnR-expressing plasmid led to restoration of its growth on DMS, DMSO and DMSO(2). An rpoN-defective mutant of strain DS1, which lacks the sigma(54) factor, grew on MSA, but not on DMS, DMSO and DMSO(2), indicating that SfnR controls expression of gene(s) involved in DMSO(2) metabolism by interaction with sigma(54)-RNA polymerase. Northern hybridization and a reporter gene assay with an sfn-lacZ transcriptional fusion elucidated that expression of the sfnECR operon was induced under sulfate limitation and was dependent on a LysR-type transcriptional regulator, CysB. This is believed to be the first report that a sigma(54)-dependent transcriptional regulator induced under sulfate limitation is involved in sulfur assimilation.
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PMID:A CysB-regulated and sigma54-dependent regulator, SfnR, is essential for dimethyl sulfone metabolism of Pseudomonas putida strain DS1. 1268 41

In Photobacterium, the flavin reductase encoded by luxG regenerates the reduced form of flavin mononucleotide (FMN). Reduced FMN is one of the substrates of the luciferase enzyme that catalyzes a light-emitting reaction. A set of experiments, that employs a luxG-expression plasmid construct (pGhis) and is suitable for an undergraduate laboratory course, is presented. Hexahistidine-tagged protein is expressed in E. coli from pGhis, with the T7 RNA polymerase/lac repressor induction system. Bacteria are lysed by sonication and the tag allows for purification by immobilized metal ion affinity chromatography. A gel filtration column is used to remove ions and the other small molecules. The Bradford assay, with multiwell plates and an automated plate reader, is used to identify protein concentration peaks from both columns. The concentration of purified enzyme is then calculated from its A(280) using the predicted extinction coefficient. Yield and purity are further assayed with SDS-PAGE. Activity of purified enzyme is measured with riboflavin or FMN as substrate. Reaction rate is quantified by monitoring decrease in A(340) as the redox partner, NADH, is oxidized.
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PMID:Expression, purification, and characterization of a recombinant flavin reductase from the luminescent marine bacterium Photobacterium leiognathi: A set of exercises for students. 2156 17