Difference between revisions of "Part:BBa K1067003"

Latest revision as of 10:27, 12 October 2013

Ammonium monooxygenase from Nitrosomonas europaea

Ammonia monooxygenase (AMO) from Nitrosomonas europaea converts ammonia to hydroxylamine;

NH₃ + O₂ + cycAX_red + 2 H⁺ → NH₂OH + H₂O + cycAX_ox

The whole operon containing amoA1, amoB1, amoC1 http://ecocyc.org/NEUR228410/NEW-IMAGE?type=MAPTICK&object=2238597/2241670&chromosome=CHROMOSOME-1&bp-range=2238597/2241670 link to Biocyc was extracted by colony PCR on a N. europaea culture.

Usage and Biology

AMO is an integral membrane protein composed of 3 subunits (amoA, amoB, amoC) and is first step in the enzymatic conversion of ammonia to nitrite in the nitrifying bacteria genus. AMO converts ammonia into the intermediate hydroxylamine. This protein works in close relationship with cycA, cycX and Hydroxylamine oxidoreductase (HAO) to complete the nitrification process by converting hydroxylamine to nitrite. HAO is providing 2 free electrons that are picked up and delivered to AMO by the cytochromes cycA and cycX, however another cytochrome from E. coli can also fill this role and the AMO can function on its own as well.

Characterization

Ammonia consumption with respect to growth (optical density) for two replicates of the AMO transformant, and untransformed E. coli control. The rate of consumption of ammonia is much higher in the transformant than in the control. (larger image)

This part is experienced to slow down growth rates of E.coli substantially; likely because this is a heterologously expressed membrane protein and thereby disrupts the E.coli membrane, and because the hydroxylamine product is toxic to E. coli.

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 263
Illegal BamHI site found at 306
Illegal BamHI site found at 2510
Illegal BamHI site found at 2822
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal AgeI site found at 2028
1000
COMPATIBLE WITH RFC[1000]

@@ Line 2: / Line 2: @@
 <partinfo>BBa_K1067003 short</partinfo>
-Ammonia monooxygenase (AMO) from ''Nitrosomonas europaea'' converts ammonia to hydroxylamine; NH<sub>3</sub> &rarr; NH<sub>2</sub>OH
+Ammonia monooxygenase (AMO) from ''Nitrosomonas europaea'' converts ammonia to hydroxylamine;
+'''NH<sub>3</sub> + O<sub>2</sub> + cycAX<sub>red</sub> + 2 H<sup>+</sup> &rarr; NH<sub>2</sub>OH + H<sub>2</sub>O + cycAX<sub>ox</sub>'''
+The whole operon containing amoA1, amoB1, amoC1 [[http://ecocyc.org/NEUR228410/NEW-IMAGE?type=MAPTICK&object=2238597/2241670&chromosome=CHROMOSOME-1&bp-range=2238597/2241670 link to Biocyc]] was extracted by colony PCR on a ''N. europaea'' culture.
-The whole operon containing amoA1, amoB1, amoC1 [http://biocyc.org/NEUR228410/NEW-IMAGE?type=OPERON-IN-CHROM-BROWSER&object=TUJNO-524 Biocyc] was extracted by colony PCR on a ''N. europaea'' culture.
-<!-- Add more about the biology of this part here
 ===Usage and Biology===
+AMO is an integral membrane protein composed of 3 subunits (amoA, amoB, amoC) and is first step in the enzymatic conversion of ammonia to nitrite in the nitrifying bacteria genus. AMO converts ammonia into the intermediate hydroxylamine.  This protein works in close relationship with [https://parts.igem.org/wiki/index.php?title=Part:BBa_K1067002 cycA], [https://parts.igem.org/wiki/index.php?title=Part:BBa_K1067002 cycX] and [https://parts.igem.org/wiki/index.php?title=Part:BBa_K1067004 Hydroxylamine oxidoreductase (HAO)] to complete the nitrification process by converting hydroxylamine to nitrite. HAO is providing 2 free electrons that are picked up and delivered to AMO by the cytochromes cycA and cycX, however another cytochrome from ''E. coli'' can also fill this role and the AMO can function on its own as well.
+===Characterization===
+https://static.igem.org/mediawiki/2013/2/25/Dtu_amo_small.png
+Ammonia consumption with respect to growth (optical density) for two replicates of the AMO transformant, and untransformed E. coli control.  The rate of consumption of ammonia is much higher in the transformant than in the control.  [https://static.igem.org/mediawiki/2013/e/ed/Dtu_Amo_confirmation.png (larger image)]
+This part is experienced to slow down growth rates of ''E.coli'' substantially; likely because this is a heterologously expressed membrane protein and thereby disrupts the ''E.coli'' membrane, and because the hydroxylamine product is toxic to ''E. coli''.
-<!-- -->
 <span class='h3bb'>Sequence and Features</span>
 <partinfo>BBa_K1067003 SequenceAndFeatures</partinfo>