Composite

Part:BBa_K2423007

Designed by: Oscar Broström   Group: iGEM17_Uppsala   (2017-10-01)
Revision as of 16:21, 31 October 2017 by Rixina (Talk | contribs)


CsADH2946 under control with BBa_J04500

This BioBrick contains the gene coding for the aldehyde dehydrogenase called CsADH2946, which is under regulation with BBa_J04500. This enzyme is a part of the second step in the zeaxanthin to crocin pathway. More specifically it catalyzes the reaction from crocetin dialdehyde to crocetin using NAD+ as a cofactor. The enzyme can be found naturally in Crocus Sativus (the plant that saffron is harvested from).

Since the gene of interest is under regulation BBa_J04500 it has to be induced with lactose or any closely related derivate such as IPTG (we used IPTG).

Usage and Biology

Saffron, a well recognized, but expensive spice has not only uses in terms of cooking but compounds found in saffron have been shown to help with inflammation (1), neurodegenerative diseases (2) and more. Some of those compounds namely zeaxanthin, crocetin dialdehyde, crocetin and crocin are all a part of the same metabolic pathway in the plant specie Crocus Sativus. Not only are these compounds in saffron helpful in terms their medicinal properties, but also the fact that they are very colorful. These aspects was what drew us at iGEM Uppsala 2017 to work with the pathway from zeaxanthin to crocin in the BioBrick format, put also to integrate the metabolic steps that leads up to crocin (the pathway from farnesyl pyrophospate (FPP) to zeaxanthin) on the chromosome of Escherichia Coli. The enzyme presented on this page catalyzes the second reaction in the zeaxanthin to crocin pathway.

In more detail CsADH2946 is an aldehyde dehydrogenase (ALDH) that oxidizes the two aldehyde groups at each end of crocetin dialdehyde to carboxylic acids. The resulting molecule from this reaction is crocetin. CsADH2946 was discovered through transcriptomic analysis of the chromoplasts of Crocus Sativus (3). The active site of CsADH2946 can be found around a loop containing three cystenin residues in a row (C337, C338, C339; positions were determined from our homology model). The residues that are conserved were found by looking at the template (PDB: 4fqf) (4) for our homology model.


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 360
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 767
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1284
    Illegal SapI.rc site found at 392


References

1. Papandreou MA, Kanakis CD, Polissiou MG, Efthimiopoulos S, Cordopatis P, Margarity M, et al. Inhibitory Activity on Amyloid-β Aggregation and Antioxidant Properties of Crocus sativus Stigmas Extract and Its Crocin Constituents. J Agric Food Chem. 2006 Nov 1;54(23):8762–8.

2. Chen L, Qi Y, Yang X. Neuroprotective effects of crocin against oxidative stress induced by ischemia/reperfusion injury in rat retina. Ophthalmic Res. 2015;54(3):157–68.

3. GĂłmez-GĂłmez L, Parra-Vega V, Rivas-Sendra A, SeguĂ­-Simarro JM, Molina RV, Pallotti C, et al. Unraveling Massive Crocins Transport and Accumulation through Proteome and Microscopy Tools during the Development of Saffron Stigma. Int J Mol Sci [Internet]. 2017 Jan 1;18(1). Available from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5297711/

4. Lang BS, Gorren ACF, Oberdorfer G, Wenzl MV, Furdui CM, Poole LB, et al. Vascular Bioactivation of Nitroglycerin by Aldehyde Dehydrogenase-2. J Biol Chem. 2012 Nov 2;287(45):38124–34.

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