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<img class="figure hundred" src="https://static.igem.org/mediawiki/2018/8/88/T--Bielefeld-CeBiTec--nanoparticles_result_LK.png"> | <img class="figure hundred" src="https://static.igem.org/mediawiki/2018/8/88/T--Bielefeld-CeBiTec--nanoparticles_result_LK.png"> | ||
<figcaption> | <figcaption> | ||
− | <b>Figure 9:</b> The silver nanoparticles in our gold silver mutant ferritin <a href="http://parts.igem.org/Part:BBa_K2638999">(BBa_K2638999)</a> with a mean diameter of 8.2 nm were significant smaller than the nanoparticles of the wildtype human ferritin (BBa_K1189019) with a mean diameter of 531.8 nm. | + | <b>Figure 9:</b> The silver nanoparticles in our gold silver mutant ferritin <a href="http://parts.igem.org/Part:BBa_K2638999">(BBa_K2638999)</a> with a mean diameter of 8.2 nm were significant smaller than the nanoparticles of the wildtype human ferritin <a href="http://parts.igem.org/Part:BBa_K1189019">(BBa_K1189019)</a> with a mean diameter of 531.8 nm. |
</figcaption> | </figcaption> | ||
</figure> | </figure> |
Revision as of 18:13, 7 December 2018
Improve a Part
Short summary
The Human Ferritin Heavy Chain (HUHF) BBa_K2638999 was successfully cloned and expressed in Escherichia coli DH5 alpha. After protein purification HUHF was used to produce gold and silver nanoparticles which was ensured by examinations with the Transmission Electron Microscope and Energy-dispersive X-ray spectroscopy (EDX). Thus, we improved BBa_K1189019 which is not able to form gold and silver nanoparticles.
The Calgary 2013 iGEM team used the human ferritin wildtype (BBa_K1189019) as reporter protein for a test strip. They expressed the human ferritin heavy and light chain heterologous using Escherichia coli. In the cells, the ferritin produced its characteristic iron core, which was colored with the help of fenton chemistry to produce the prussian blue iron complex. Beside the function as reporter, the team mentioned the capability of ferritin to produce nanoparticles from other metal ions.
Improved Human Ferritin: BBa_K2638999
Figure 7 shows a TEM image with 147 identified silver nanoparticles produced by the wild type human ferritin (BBa_K1189019). The particles are between 24.5 and 1597.8 nm in size with one very big particle with a size of 7272.3 nm, which seems to consist in many agglutinated silver nanoparticles. No particle was found in the expected size of about 8 nm.
Figure 8 shows a TEM image with 708 identified silver nanoparticles produced by the gold silver mutant ferritin sample (BBa_K2638999). The particles have a size between 1.8 and 34.8 nm. 120 of the silver nanoparticles (16.9 %) are exactly in the expected size of 7 to 9 nm which indicates that at least all of these particles are produced by our improved ferritin (BBa_K2638999).
The direct comparison of our new gold silver mutant ferritin (BBa_K2638999) and the old wild type human ferritin (BBa_K1189019) in figure 9 shows that our improved enzyme produces nearly five times more silver nanoparticles which are 98.5 % smaller than the silver nanoparticles produced by the wild type ferritin. This proves that the new ferritin enzyme is much more suitable for producing silver nanoparticles than the wild type version.
Figure 10 shows two gold nanoparticles of 13 and 10 nm diameter that were produced by the wild type human ferritin sample (BBa_K1189019). They are slightly bigger than the expected size between 7 and 9 nm, and thus it cannot be ensured that these particles are really produced by that enzyme.
Figure 11 shows two gold nanoparticles of 7 and 9 nm diameter that were produced by the gold silver mutant ferritin (BBa_K2638999). They are exactly in the expected size range, although it is difficult to draw reliable conclusions from this small size and number of particles.
Outlook
Molecular graphics and analyses performed with UCSF Chimera, developed by the Resource for Biocomputing, Visualization, and Informatics at the University of California, San Francisco, with support from NIH P41-GM103311.
Butts, C.A., Swift, J., Kang, S., Di Costanzo, L., Christianson, D.W., Saven, J.G., and Dmochowski, I.J. (2008).. Directing Noble Metal Ion Chemistry within a Designed Ferritin Protein † , ‡. Biochemistry 47: 12729–12739.
Castro, L., Blázquez, M.L., Muñoz, J., González, F., and Ballester, A. (2014).. Mechanism and Applications of Metal Nanoparticles Prepared by Bio-Mediated Process. Rev. Adv. Sci. Eng. 3.
Ensign, D., Young, M., and Douglas, T. (2004).. Photocatalytic synthesis of copper colloids from CuII by the ferrihydrite core of ferritin. Inorg. Chem. 43: 3441–3446.
Goujon, M., McWilliam, H., Li, W., Valentin, F., Squizzato, S., Paern, J., and Lopez, R. (2010).. A new bioinformatics analysis tools framework at EMBL-EBI. Nucleic Acids Res. 38: W695-699.
Pettersen, E.F., Goddard, T.D., Huang, C.C., Couch, G.S., Greenblatt, D.M., Meng, E.C., and Ferrin, T.E. (2004).UCSF Chimera--a visualization system for exploratory research and analysis. J Comput Chem 25: 1605–1612.
Sievers, F., Wilm, A., Dineen, D., Gibson, T.J., Karplus, K., Li, W., Lopez, R., McWilliam, H., Remmert, M., Söding, J., Thompson, J.D., and Higgins, D.G. (2011). Fast, scalable generation of high-quality protein multiple sequence alignments using Clustal Omega. Mol. Syst. Biol. 7: 539.
Ummartyotin, S., Bunnak, N., Juntaro, J., Sain, M., and Manuspiya, H. (2012). . DSynthesis of colloidal silver nanoparticles for printed electronics. /data/revues/16310748/v15i6/S1631074812000549/.
Wang, L., Hu, C., and Shao, L. (2017a).. The antimicrobial activity of nanoparticles: present situation and prospects for the future. Int. J. Nanomedicine 12: 1227–1249.
Wang, Z., Gao, H., Zhang, Y., Liu, G., Niu, G., and Chen, X. (2017b).. Functional ferritin nanoparticles for biomedical applications. Front. Chem. Sci. Eng. 11: 633–646.