Coding
MaoA

Part:BBa_K1615022

Designed by: Brooke Rothschild-Mancinelli   Group: iGEM15_Edinburgh   (2015-08-28)
Revision as of 06:39, 19 October 2021 by Anthneedee (Talk | contribs)

Monoamine Oxidase A RFC25

Monoamine oxidase A is encoded by the gene maoA and is subject to catabolite and ammonium ion repression1. Amine oxidases that contain copper/topaquinone (TPQ), like monoamine oxidase A, convert primary amines into their corresponding aldehydes, hydrogen peroxide and ammonia2.

To test the activity of monoamine oxidase A, tyramine can be used as a substrate and its corresponding aldehyde as well as ammonia and hydrogen peroxide will be produced. When the hydrogen peroxide is coupled with horseradish peroxidase and Amplex red, resorufin is produced which has a strong red colour.

1Oka, M., Murooka, Y., & Harada, T. (1980). Genetic control of tyramine oxidase, which is involved in derepressed synthesis of arylsulfatase in Klebsiella aerogenes. Journal of bacteriology, 143(1), 321-327.
2McIntire, W. S., & Hartmann, C. (1993). Copper-containing amine oxidases. Principles and applications of quinoproteins, 97-171.
3Sugino, H., Sasaki, M., Azakami, H., Yamashita, M., & Murooka, Y. (1992). A monoamine-regulated Klebsiella aerogenes operon containing the monoamine oxidase structural gene (maoA) and the maoC gene. Journal of bacteriology, 174(8), 2485-2492.


Contribution (Waterloo iGEM 2021)

Summary: The role of monoamine oxidase A in the metabolism of biogenic amines, including neurotransmitters implicated in ADHD, made it a potential candidate biomarker for Waterloo iGEM 2021's ADHD diagnosis tool, NeuroDetech. Ultimately, monoamine oxidase was not utilized in our final design. However, to inform this decision, we conducted literature research into the structure, function, and stability of monoamine oxidase A. We felt that our findings would be effective in supplementing the existing documentation on this part; thus, our findings are discussed below.

Documentation:

Monoamine oxidase is a homodimer that requires a number of cofactors for proper functioning, including the following:

  • Copper(II) ions are a main regulator of activity. For this reason, monoamine oxidase is considered a copper amine oxidase (Wilmot et al., 1997).
  • 4 calcium(II) ions; 2 calcium(II) ions bind to each subunit of monoamine oxidase (Wilmot et al., 1997).
  • Topaquinone, a derivative of tyrosine. Topaquinone is associated with the copper amine oxidase activity of monoamine oxidase A, so it presence is required in addition to copper(II). With that said, the presence of copper(II) ions facilitates the conversion of tyrosine to topaquinone (Murray et al., 1999).

In addition to tyramine (as described by iGEM Edinburgh in the documentation above), other known substrates of monoamine oxidase include 3-nitrotyramine and phenylethylamine, the latter of which is an ADHD-associated neurotransmitter (Rankin et al., 2008).

Waterloo iGEM's 2021 project, NeuroDetech, detects biomarkers in urine samples. When initially considering monoamine oxidase as a biomarker, we were concerned about its stability in urine. Thus, we reviewed the stability of monoamine oxidase under the parameters of temperature and pH, as discussed below:

  • Temperature stability: Elovaara et al. (2015) measured the activity of monoamine oxidase using assays conducted at 37 C to replicate human body temperature. No inhibition of the enzyme was reported, and thus human urine temperature was not deemed to cause denaturing or inhibition of the enzyme (Elovaara et al., 2015)
  • pH stability: Murakawa et al. (2015) probed the mechanism of a monoamine oxidase from Arthrobacter species. In their methods, they used pH values ranging from 5.7-6.7. Alternatively, Elovaara et al. (2015) assayed monoamine oxidase where the most common pH value was 7.0, while Kurtis et al. (2011) used a pH of 6.0 in their methods. Overall, a slightly acidic to neutral pH range seemed to be suitable for proper activity of monoamine oxidase.

The following is the 3D structure of monoamine oxidase A in its homodimeric form. The 3D structure was obtained using UCSF Chimera (Pettersen et al., 2004).


References:

Elovaara, H., Huusko, T., Maksimow, M., Elima, K., Yegutkin, G. G., Skurnik, M., Dobrindt, U., Siitonen, A., McPherson, M. J., Salmi, M., & Jalkanen, S. (2015, November 10). Primary amine oxidase of Escherichia coli is a metabolic enzyme that can use a human leukocyte molecule as a substrate. PLoS One, 10(11), e0142367. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4640556/

Kurtis, C. R. P., Knowles, P. F., Parsons, M. R., Gaule, T. G., Phillips, S. E. V., & McPherson, M. J. (2011, March 10). Tyrosine 381 in E. coli copper amine oxidase influences substrate specificity. Journal of Neural Transmission, 118, 1043–1053. https://link.springer.com/article/10.1007/s00702-011-0620-y

Murakawa, T., Hamaguchi, A., Nakanishi, S., Kataoka, M., Nakai, T., Kawano, Y., Yamaguchi, H., Hayashi, H., Tanizawa, K., & Okajima, T. (2015). Probing the Catalytic Mechanism of Copper Amine Oxidase from Arthrobacter globiformis with Halide Ions. Enzymology, 290(38), P23094-23109. https://www.jbc.org/article/S0021-9258(20)44762-3/fulltext

Murray, J. M, Saysell, C. G., Wilmot, C. M., Tambyrajah, W. S., Jaeger, J., Knowles, P. F., Phillips, S. E. V., & McPherson, M. J. (1999). The Active Site Base Controls Cofactor Reactivity in Escherichia coli Amine Oxidase:  X-ray Crystallographic Studies with Mutational Variants. Biochemistry, 38(26), 8217–8227. https://pubs.acs.org/doi/10.1021/bi9900469

Pettersen, E. F.; Goddard, T. D.; Huang, C. C.; Couch, G. S.; Greenblatt, D. M.; Meng, E. C. & Ferrin, T. E. UCSF Chimera--a visualization system for exploratory research and analysis (Version 1.15). J Comput Chem. 2004; 25(13): 1605-1612. https://www.ncbi.nlm.nih.gov/pubmed/15264254

Rankin, L. D., Bodenmiller, D. M., Partridge, J. D., Nishino, S. F., Spain, J. C., Spiro, S. (2008, September 15). Escherichia coli NSRR regulates a pathway for the oxidation of 3-nitrotyramine to 4-Hydroxy-3-nitrophenylacetate. Journal of Bacteriology, 190(18), 6170-7. https://journals.asm.org/doi/10.1128/JB.00508-08.

Wilmot, C. M., Murray, J. M., Alton, G., Parsons, M. R., Convery, M. A., Blakeley, V., Corner, A. S., Palcic, M. M., Knowles, P. F., McPherson, M. J., & Phillips, S. E. V. (1997). Catalytic Mechanism of the Quinoenzyme Amine Oxidase from Escherichia coli:  Exploring the Reductive Half-Reaction. Biochemistry, 36(7), 1608–1620. https://pubs.acs.org/doi/10.1021/bi962205j


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


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