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URL: http://www.addgene.org/78201
Proper Citation: RRID:Addgene_78201
Insert Name: parM (K33A I27C T174A T175N C287A)
Organism: E. coli
Bacterial Resistance: Ampicillin
Defining Citation: PMID:19801632
Vector Backbone Description: Vector Backbone:pHis17; Vector Types:Bacterial Expression; Bacterial Resistance:Ampicillin
Comments: The plasmid is used to express the plasmid segregation protein, ParM, from E. col. The protein is modified for use as the scaffold of an ADP biosensor, MDCC-ParM. A single cysteine mutation is for label attachment (I27C); there is a mutation to inhibit filament formation (K33A), two mutations to weaken ATP binding (T174A and T175N), a mutation to change the exposed cysteine (C287A) and a C-terminal His6 tag to aid purification. When labelled with a single N-[2-(1-maleimidyl)ethyl]-7-diethylaminocoumarin-3-carboxamide (MDCC) fluorophore, this biosensor has a fluorescence intensity change of up to 4-fold and can be used to measure ADP in the range of tens of nanomolar to tens of micromolar in the presence of up to~200 μM ATP.
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Source: Addgene
