Team:Hong Kong-CUHK/Description

The Magnetosome

Magnetosome is a membrane bounded intracellular structure, something that is actually rare in prokaryotic organisms like bacteria. They are of nano-size ranging from about 35-120 nm. Magnetosomes comprise of a magnetic mineral crystal surrounded by a lipid bilayer membrane about 3–4 nm thick (fig. _1_). A number of common cytoplasmic membrane fatty acids components can be found in the magnetosome membrane. It is then later highly suspected that magnetosomes are membrane invaginations originating from the cytoplasmic membrane [1]. The magnetosome membrane is highly significant as it creates an isolated environment within the cell which is crucial for mineral crystal nucleation and growth [2]. These membrane-enclosed inorganic crystals consist of either the magnetic minerals magnetite (Fe3O4) or greigite (Fe3S4). The particles usually arrange themselves along the cell axis either in one or multiple chains . Different varieties of crystal morphologies such as cubo-octahedral, elongated hexagonal prismatic, and bullet-shaped morphologies have been discovered [1]

.

Figure 1: Magnetosome

The Magnetotactic Bacteria -- The origin of magnetosome

Magnetosomes are intracellular membrane bounded organelle synthesized by Magnetotactic bacteria. First discovered in 1975 by Richard Blakemore, these magnetotactic bacteria are mobile, aquatic, gram-negative prokaryotes [3] with a myriad of cellular morphologies, including coccoid, rod-shaped, vibrioid, helical or even multi-cellular. They are found in their highest numbers at, or just below, the oxic-anoxic interface in aquatic habitats and exhibit a negative growth response to atmospheric concentrations of oxygen.

Magnetosomes form a chain and align themselves along the axis within the bacteria. With the formation of magnetosomes inside them, they are able to align passively to the earth’s magnetic field and hence use minimum energy to swim along geomagnetic field lines. This behaviour is called magnetotaxis [4] and is beneficial to the survival of the bacteria as it helps them to reach regions of optimal oxygen concentrations without random, unnecessary movements [5].

Why and What is Azotobacter?

Though magnetotactic bacteria is the origin of magnetosomes, these bacteria are described by scientists as a group of fastidious prokaryotic bacteria -- meaning that they are difficult to cultivate owing to their unusual growth requirements. Their micro-aerophilic nature require elaborate growth techniques, and they are difficult to grow on the surface of agar plates, which would make the screening for mutants a problem [6].

The lack of effective methods of DNA transfer in these microorganisms is a challenge too. Luckily, the situation is improving due to better technologies recently and some of the genes from M. magnetotacticum have been confirmed functionally expressed in E. coli. This shows that the transcriptional and translational elements of the two microorganisms are compatible. With such good news, a number of previous igem teams including kyoto: 2014, OCU-China: 2013, Washington: 2011 and UNIK_Copenhagen: 2013 have been working with transferring the magnetosome related genes to e.coli. Some exciting results the formation of magnetosome membrane in e.coli (by the kyoto team) has been reported by previous teams, however, never the whole magnetosome. We have been wondering why magnetosomes seems so hard to be formed in e.coli. And then, we come up with a hypothesis -- the formation of magnetosome requires a micro-aerobic or anaerobic environment as the magnetotatic bacteria are all living micro-aerobically.

Therefore, we chose a new bacteria to work on our magnetosome project -- the Azotobacter vinelandii. Azotobacter vinelandii is gram-negative diazotroph. It is a soil bacterium related to the Pseudomonas genus that fixes nitrogen under aerobic conditions while having enzymatic mechanisms protecting its oxygen-sensitive nitrogenase from oxygen damage. This findings shows that A. vinelandii is an excellent host for the production and characterization of oxygen-sensitive proteins or organelles as in our case [7].

With the biggest advantage of using azotobacter is that it is an aerobic bacteria with an intracellular anaerobic condition, we can grow it easily in normal conditions in lab without expensive equipments like fermenter, while fulfilling the growing criteria for the magnetosome. Besides, most parts in registry are functionable in Azotobacter and Azotobacter is of safety level group 1 too. One more important thing is that it can do homologous recombination by itself which is a critical process we need in our project.