Aconitase Enzyme Activity Microplate Assay Kit

Catalog No. MS745

$425.00 - 96 tests


Product Description

Kit MS745 measures the activity of aconitase in a sample by following the conversion of isocitrate to cis-aconitate as an increase in absorbance at UV 240 nm with an extinction coefficient of 2.2 OD/mM per well. Aconitase preservation solution, assay buffer, reagents and a special UV plate are provided for this measurement by a UV 240nm capable spectrophotometer.

Aconitase is an iron-sulfur protein that catalyzes the reversible interconversion of citrate and isocitrate, via a cis-aconitate intermediate, in both the TCA and glyoxylate cycles. The enzyme contains a [4Fe-4S] cluster which interacts directly with the substrates. In eukaryotes there are mitochondrial (ACO2) and cytosolic (ACO1) forms of the enzyme. The mitochondrial form functions not only in the TCA cycle, but also to stabilize mtDNA thereby influencing mitochondrial gene expression. The cytosolic form can function as an aconitase as well as an iron regulatory protein. Both forms are sensitive to oxidants, which can inactivate the enzyme by changing the [4Fe-4S] to a [3Fe-4S] cluster. This change in mitchondrial aconitase can lead to decreased energy production, whereas in cytosolic aconitase it triggers binding of the enzyme to mRNA iron response elements resulting in increased expression of iron uptake proteins and decreased transcription of iron sequestering protein.

An inactivated [3Fe-4FS] aconitase may be activated in vitro by the addition of iron and cysteine. The active form of the enzyme is inhibited by citrate analogs, fluoracetate, and redox stress agents. Other inhibitors include oxidative stress agents such as peroxynitrite, hydrogen peroxide and superoxide, which lead to [3Fe-4FS] inactivation of the enzyme. Therefore, aconitase is considered a good marker of mitochondrial and cellular oxidative stress.



 
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Product Specifications
 
Species Reactivity: Human, mouse, rat, bovine
Storage Conditions: Buffer, detergent, and microplate should be stored at 4°C.
Reagent C (reduced cytochrome c) should be stored at -20°C.
Country of Origin: USA


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Figure 1



(click to enlarge)

Figure 1. Bovine heart mitochondria were treated with increasing concentrations of hydrogen peroxide and peroxynitrite to generate aconitase activity IC50's for these oxidative stress agents.
Figure 2



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Figure 2. Using Mitosciences’ cellular fractionation kit MS861, whole cells were separated into cytoplasmic, mitochondrial and nuclear fractions.


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Downloadable Documents


   Technical Data Sheet

   Protocol


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