Difference between revisions of "Part:BBa K2346000"

Revision as of 09:16, 30 October 2017

Nitric oxide reductase (NOR)

Cytochrome P450 55B1 from Chlamydomonas reinhardtii is reported to function as a nitric oxide reductase (NOR). We are still working on that. Since nitric oxide is nearly insoluable in water, it’s hard for us to create a aquatic reaction system for assaying the enzyme reactivity. We are still working on that. But currently we plan to create a reaction environment in a sealed container filled with solution, enzyme, and nitric oxide gas, By constant shake over, let’s say, two hours, we’ll then measure the change in the concentration of NO using the Gas-Phase Molecular Absorption Spectrometry to see if the presence of the enzyme made any difference on the concentration.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 1
Illegal XhoI site found at 1216
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal AgeI site found at 1200
1000
COMPATIBLE WITH RFC[1000]

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 <partinfo>BBa_K2346000 short</partinfo>
-From Agrobacterium tumefaciens. The enzyme contains two copper center, namely CuA and Cuz, which can transfer the electron.
+Cytochrome P450 55B1 from Chlamydomonas reinhardtii is reported to function as a nitric oxide reductase (NOR). We are still working on that. Since nitric oxide is nearly insoluable in water, it’s hard for us to create a aquatic reaction system for assaying the enzyme reactivity. We are still working on that. But currently we plan to create a reaction environment in a sealed container filled with solution, enzyme, and nitric oxide gas, By constant shake over, let’s say, two hours, we’ll then measure the change in the concentration of NO using the Gas-Phase Molecular Absorption Spectrometry to see if the presence of the enzyme made any difference on the concentration.
-The assay:
-We prepared a standard measuring mixture(2ml) consisting of 50mM Tris-HCL,pH9, 0.5mM methyl viologen, a proper amount of NosZ and 1mM sodium dithionite. Methyl viologen and sodium dithionite will go on to react and form a blue intermediate product, with a relativelly stable absorbance at 600nm. The blue interdiate product actually serves as an electron donor for the reduction of nitrous oxide. Therefore, as the reaction goes on, the blue substance starts to lose electrons and its color dims, resulting in a lower 600nm absorbance. Thus, by directly measuring the change of color in the system, we are able to track the processing of the reaction.