Team:ZJU-China/Magnetosome
Magnetosome
Considering that magnetotactic bacteria such as Magnetospirillum gryphiswaldense MSR-1 is not a common bacteria that known by major scholars. In this page, we will give a brief introduction about magnetotactic bacteria and magnetosomes which related to our project to help readers understanding our idea better.
Magnetotactic Bacteria
Magnetotactic bacteria, were discovered by Blakemore in 1975. The characteristic of the bacterium is magnetotactic due to its cell containing magnetosomes consisting of iron crystals that often arrange in chains along the magnetic lines. The iron crystals are made of Fe3O4, Fe3S4, Fe2O3, FeS etc[1], and are enveloped by the cytoplasmic membrane. The first strain of pure culture of this kind of bacterium was Aquaspirillum magnetotacticum, Magnetospirillum gryphiswaldense MSR-1 and M. magnetotacticum MS-1 has been isolated and identified one after another.
Magnetotactic bacteria might serve as a model system for understanding the physiological function of magnetic crystals in higher organisms and even cerebrum of human [2].
Formation of Magnetosome
The formation of magnetosomes in magnetotactic bacteria is called biomineralization, of which the specific mechanisms are still unclear at present, however. By genetic methods such as T-DNA insertion, researchers construct different loss-of-function mutants of several species of magnetotactic bacteria to screen out the genes that control biomineralizations. As yet, more than 40 different genes have been identified that encode magnetosome-associated proteins and/or that yield magnetosomes-related phenotypes in AMB-1 and MSR-1 mutants. Although several auxiliary, non-essential functions are contributed by general metabolic and regulatory cellular pathways that are not specific to magnetosome biogenesis, all specific functions in magnetosomes biogenesis are associated with a set of about 30 mam (magnetosome membrane) and mms (magnetic particle-membrane specific) genes that are clustered in a single chromosomal region, the genomic magnetosome island. The magnetosome island extends across about 100kb, which is equivalent to approximately 2% of a typical MSR-1 genome and shares characteristics with other bacterial genomic island (such as putative insertion sites near tRNA genes, distinct levels of GC enrichment, and a large number of mobile genetic elements)[3][4][5].
The formation of the magnetosomes can be divided into 4 main steps[6].
1.Formation of magnetosome membrane. Compartmentalization of magnetosome biogenesis by the magnetosome membrane provides the growing magnetite crystal with a specialized ‘nano-reactor’ in which the iron, redox and pH environments of bio‑mineralization can be strictly regulated. The magnetosome membrane was shown to comprise a nm proteinaceous phospholipid bilayer that is invaginated from the cytoplasmic membrane. The presence and interaction of several magnetosome proteins are required to induce membrane curvature and vesicle formation. It has been suggested that MamB might be a landmark protein for the formation of protein interaction networks that induce membrane curvature.
2.Targeting proteins to the magnetosome membrane. This step is tither before, coincidently or following the invagination of magnetosome membrane. All Mam and Mms proteins that have been studied in detail have been found to associate with purified magnetosomes, though the quantity that is present in purified magnetosome preparations varies for each other. Furthermore, localization to the magnetosome membrane has been confirmed for many of these proteins by immunolabelling or fluorescence tagging in intact cells, however, how proteins are specifically targeted to the magnetosome membrane is not currently known, and no motifs that encode sorting signals to the magnetosome membrane have been identified.
3.Biomineralization of magnetite crystals. Following the formation of magnetosome membrane, a process of biomineralization occurs in which large amounts of iron are accumulated in magnetosome membrane vesicles. Currently, most evidence supports the notion that iron accumulation in magnetosome membrane vesicles occurs subsequently to iron uptake by the cell. For example, in MSR, loos of the GTP-dependent Fe2+ transporters FeoB1 and FoeB2, which are probably distributed over the entire cytoplasmic membrane and not specific to the entire magnetosome membrane, caused a strong impairment of biomineralization. Then, the absorbed iron ions are utilized to grow magnetite within a narrow redox range as it is mixed-valence iron oxide. Nucleation of magnetite crystals occurs when, under optimal conditions (>pH 7 and low redox potential), soluble Fe2+ and Fe3+ start to form crystalline iron phases.
4.Assembly of magnetosome chains. Magnetosomes are aligned into linear chains in which magnetic moments of individual particles sum up to maximize the total magnetic responses of the cell. Depending on the growth conditions, magnetosome chains in MSR-1 can contain more than 100 magnetosomes. Magnetosome chain assembly results from the coordinated interplay of magnetic interactions, which occur naturally between magnetite crystals, and active assembly mechanisms, which are mediated by proteins. Cryoelectron tomograms of MSR and AMB revealed the existence of bundles of cytoskeletal filaments that traverse the cell in close proximity to magnetosome chains. These filaments are formed by MamK, which is an actin-like protein that is mostly restricted to magnetotactic bacteria, in which it is highly conserved, and encoded by a gene in the mamAB operon. Rather than simply providing a rigid scaffold for chain assembly, MamK has been suggested to have an active role in the process that may involve the generation of mechanical forces through polymerization and depolymerization, similarly to other cytomotive cytoskeletal filaments.
Each step is regulated by magnetosome island precisely, and the difference between different species of magnetotactic bacteria are the shape of crystals they produce[8][9][10][11].Among the proteins that encoded by magnetosome island, MamC, which is also named Mms13, though is not a key protein that regulates the 4 steps above, yet covers most area on the surface of the magnetosome membrane. Details of the loading process of MamC is still unclear. Structure-function studies showed that the MamC has 2 transmembrane domains, leading its N- and C-terminal are both localized to the cytoplasmic side. On the inner side of the magnetosomes, MamC has a magnetite-interaction loop structure that connect the Fe3O4 directly to regulate the shape of the magnetite crystals [13][14]. By taking advantages of these features, MamC has the potential to be an anchor molecule to link nucleic acid and proteins by surface display technology[15].
Current Shortcoming of MRI
The existing contrast agents for MRI are common-used for all kinds of cancers, namely, they are
Magnetosome for MRI
Magnetotactic bacteria is a general term for a group of bacteria which is gram-negative bacteria
that can move along magnetic field under the action of an external magnetic
field due to a series of magnetite crystals designated as magnetosomes in their cytoplasm[16]. Each magnetosome consists of an internal
magnetite crystal and outer bilayer
phospholipid membranes. Dozens of magnetosomes are arranged in chains within magnetotactic
bacteria by the action of cytoskeleton. Magnetosomes have the characteristics of
nanometer-size, single magnetic domain, large specific surface area, good biocompatibility,
superparamagnetism and so on, especially they can be modified through genetic
engineering methods, thus magnetosomes are widely used in medical field.
Compared with artificially synthesized nanosized magnetite, it is the most impressive that
magnetosomes have naturally formed phospholipid bilayer membrane. Besides that,
magnetosomes have the characteristics of stable single magnetic domain structure, permanent
magnetism at room temperature, high chemical purity and uniform particle size and
crystal shape[17]. Taking advantages of these features, we
can construct magnetosomes modified with target molecular such as antibodies on
its surface to obtain specific contrast agent for different diseases.
Specificity Demonstration
Recently, the potential application of magnetosomes as MRI contrast agent has been confirmed in several cell models, including pancreatic cells, brain cells and xenograft tumor cells[18][19]. Surface-modified magnetosomes with a novel peptide (P75) targeting HER2 specifically bind to cancer cells as well as xenograft tumors with surprisingly low accumulation in other organs in vitro and vivo fluorescence imaging results and show obviously negative contrast enhancement in vivo T2-weighted MR imaging[20], which provides better imaging performance and specificity than existing contrast agents. Different to their design, we chose scFv as our targeting molecular to the HER2-positive tumor cells. The scFv may cause less or even no immune responses. Besides that, based on the diversity of scFv, we can build a platform to design relative antibodies that target different tumors.
References
[1]. Jiang, W. , Zhao, D. , Li, Y. , Tian, J. , Wang, Z. , & Li, J. . (2002). Submerged culture of magnetospirillum gryphiswaldense under n2-fixing condition and regulation of activity of nitrogen fixation. Chinese Science Bulletin, 47(24), 2095-2099
[2]. Kirschvink, J. L., Kobayashi-Kirschvink, A., & Woodford, B. J. (1992). Magnetite biomineralization in the human brain. Proceedings of the National Academy of Sciences of the United States of America, 89(16), 7683–7687. https://doi.org/10.1073/pnas.89.16.7683
[3]. Liu, J., Zhang, W., Li, X., Li, X., Chen, X., Li, J. H., Teng, Z., Xu, C., Santini, C. L., Zhao, L., Zhao, Y., Zhang, H., Zhang, W. J., Xu, K., Li, C., Pan, Y., Xiao, T., Pan, H., & Wu, L. F. (2017). Bacterial community structure and novel species of magnetotactic bacteria in sediments from a seamount in the Mariana volcanic arc. Scientific reports, 7(1), 17964. https://doi.org/10.1038/s41598-017-17445-4
[4]. Richter, M., Kube, M., Bazylinski, D. A., Lombardot, T., Glöckner, F. O., Reinhardt, R., & Schüler, D. (2007). Comparative genome analysis of four magnetotactic bacteria reveals a complex set of group-specific genes implicated in magnetosome biomineralization and function. Journal of bacteriology, 189(13), 4899–4910. https://doi.org/10.1128/JB.00119-07
[5]. Ullrich, S., Kube, M., Schübbe, S., Reinhardt, R., & Schüler, D. (2005). A hypervariable 130-kilobase genomic region of Magnetospirillum gryphiswaldense comprises a magnetosome island which undergoes frequent rearrangements during stationary growth. Journal of bacteriology, 187(21), 7176–7184. https://doi.org/10.1128/JB.187.21.7176-7184.2005
[6]. Dobrindt, U., Hochhut, B., Hentschel, U., & Hacker, J. (2004). Genomic islands in pathogenic and environmental microorganisms. Nature reviews. Microbiology, 2(5), 414–424. https://doi.org/10.1038/nrmicro884
[7]. Uebe, R., & Schüler, D. (2016). Magnetosome biogenesis in magnetotactic bacteria. Nature reviews. Microbiology, 14(10), 621–637. https://doi.org/10.1038/nrmicro.2016.99
[8]. Nies D. H. (2011). How iron is transported into magnetosomes. Molecular microbiology, 82(4), 792–796. https://doi.org/10.1111/j.1365-2958.2011.07864.
[9]. Lefèvre, C. T., Trubitsyn, D., Abreu, F., Kolinko, S., Jogler, C., de Almeida, L. G., de Vasconcelos, A. T., Kube, M., Reinhardt, R., Lins, U., Pignol, D., Schüler, D., Bazylinski, D. A., & Ginet, N. (2013). Comparative genomic analysis of magnetotactic bacteria from the Deltaproteobacteria provides new insights into magnetite and greigite magnetosome genes required for magnetotaxis. Environmental microbiology, 15(10), 2712–2735. https://doi.org/10.1111/1462-2920.12128
[10]. Kolinko, S., Richter, M., Glöckner, F. O., Brachmann, A., & Schüler, D. (2016). Single-cell genomics of uncultivated deep-branching magnetotactic bacteria reveals a conserved set of magnetosome genes. Environmental microbiology, 18(1), 21–37. https://doi.org/10.1111/1462-2920.12907
[11]. Lefèvre, C. T., & Bazylinski, D. A. (2013). Ecology, diversity, and evolution of magnetotactic bacteria. Microbiology and molecular biology reviews : MMBR, 77(3), 497–526. https://doi.org/10.1128/MMBR.00021-13
[12]. Uebe, R., & Schüler, D. (2016). Magnetosome biogenesis in magnetotactic bacteria. Nature reviews. Microbiology, 14(10), 621–637. https://doi.org/10.1038/nrmicro.2016.99
[13]. Quinlan, A., Murat, D., Vali, H., & Komeili, A. (2011). The HtrA/DegP family protease MamE is a bifunctional protein with roles in magnetosome protein localization and magnetite biomineralization. Molecular microbiology, 80(4), 1075–1087. https://doi.org/10.1111/j.1365-2958.2011.07631.x
[14]. Nudelman, H., Valverde-Tercedor, C., Kolusheva, S., Perez Gonzalez, T., Widdrat, M., Grimberg, N., Levi, H., Nelkenbaum, O., Davidov, G., Faivre, D., Jimenez-Lopez, C., & Zarivach, R. (2016). Structure-function studies of the magnetite-biomineralizing magnetosome-associated protein MamC. Journal of structural biology, 194(3), 244–252. https://doi.org/10.1016/j.jsb.2016.03.001
[15]. Nudelman, H., Perez Gonzalez, T., Kolushiva, S., Widdrat, M., Reichel, V., Peigneux, A., Davidov, G., Bitton, R., Faivre, D., Jimenez-Lopez, C., & Zarivach, R. (2018). The importance of the helical structure of a MamC-derived magnetite-interacting peptide for its function in magnetite formation. Acta crystallographica. Section D, Structural biology, 74(Pt 1), 10–20. https://doi.org/10.1107/S2059798317017491
[16]. Lefèvre, C. T., & Bazylinski, D. A. (2013). Ecology, diversity, and evolution of magnetotactic bacteria. Microbiology and molecular biology reviews : MMBR, 77(3), 497–526. https://doi.org/10.1128/MMBR.00021-13
[17]. Uebe, R., & Schüler, D. (2016). Magnetosome biogenesis in magnetotactic bacteria. Nature reviews. Microbiology, 14(10), 621–637. https://doi.org/10.1038/nrmicro.2016.99
[18]. Lee, N., Kim, H., Choi, S. H., Park, M., Kim, D., Kim, H. C., Choi, Y., Lin, S., Kim, B. H., Jung, H. S., Kim, H., Park, K. S., Moon, W. K., & Hyeon, T. (2011). Magnetosome-like ferrimagnetic iron oxide nanocubes for highly sensitive MRI of single cells and transplanted pancreatic islets. Proceedings of the National Academy of Sciences of the United States of America, 108(7), 2662–2667. https://doi.org/10.1073/pnas.1016409108
[19]. Boucher, M., Geffroy, F., Prévéral, S., Bellanger, L., Selingue, E., Adryanczyk-Perrier, G., Péan, M., Lefèvre, C. T., Pignol, D., Ginet, N., & Mériaux, S. (2017). Genetically tailored magnetosomes used as MRI probe for molecular imaging of brain tumor. Biomaterials, 121, 167–178. https://doi.org/10.1016/j.biomaterials.2016.12.013
[20]. Xiang, Z., Yang, X., Xu, J., Lai, W., Wang, Z., Hu, Z., Tian, J., Geng, L., & Fang, Q. (2017). Tumor detection using magnetosome nanoparticles functionalized with a newly screened EGFR/HER2 targeting peptide. Biomaterials, 115, 53–64. https://doi.org/10.1016/j.biomaterials.2016.11.022
