Meet Magneto Anew: Evolving Enhanced Magnetism in Bacteria

Maliha Tanjum Chowdhury
Sophomore
School of Life Sciences
Independent University, Bangladesh

November 3, 2017

Magneto. A mutant arisen during the tyranny of Nazi rule – an abominable villain who possesses the power to wield and crumple all that is metal-made, vowing to crush anything and anyone obstructing his path. There stands not a single X-Men fan who hasn’t been mesmerized and equally appalled by the terror this mutant is capable of. This article, too, discusses mutants with a special affinity for metals (more specifically, iron). However, they are thankfully much more benign than our infamous bad-boy Magneto. I’m talking about bacterial mutants that have enhanced magnetism in their iron-binding protein. My fellow geeks, this may not be as groundbreaking as a human being who is able to spontaneously generate magnetic fields, but the fact that a natural protein present in bacteria has been selectively evolved to bring about considerable magnetic properties is in itself pretty incredible. As we will later discuss, these magnetic bacteria could have important applications in research and medicine. Now sit back, read on and let your minds be blown.

The paper I am about to break down is heftily titled, “Engineering Genetically-encoded Mineralization and Magnetism via Directed Evolution.” The subject of experimentation was the natural iron-sequestering protein ferritin in the bacteria E. coli (versions of the protein are found widely in all domains of life, including humans). This protein mainly serves as a homeostatic regulator of cellular free-iron levels, storing iron to be released in times of need (incorporating free iron into a cellular protein is an example of biomineralization). Ferritin stores iron in a hydrated, amorphous form of iron oxide, which is a biocompatible substance (harmonious with living tissue), and is known to be magnetic when crystalline. The mineralized iron stored in ferritin displays poor crystallinity, and thus has a very low magnetic moment, which confers it with the ability to release iron with sufficient ease in times of deficiency. Though the factors affecting crystallinity and hence magnetic properties of the iron oxide core within ferritin remain largely unclear, this protein was deemed a model starting point for this study which sought to find ways of increasing the innate magnetism of the core by changing the structure of the protein. As an aside, magnetic bacteria do exist in nature (magnetotactic bacteria), but here they were building magnetic versions of a normally non- or poorly magnetic protein.

The mutant ferritin-containing bacteria (H34+T64I) demonstrably concentrate around magnets

The scientists first needed a system to assess iron uptake by ferritin. The problem was that cells have other means of dealing with any iron that is provided to them, such as other iron-storage proteins and metal cation exporters. To solely observe iron-sequestration by ferritin, the scientists created special “knock-out” strains of the bacteria that had all genes that interact with iron deleted. They could then add mutant ferritin genes to these bacteria via plasmids, and measure iron uptake or select mutants with greater ability to bind iron. Plasmids are circular pieces of DNA that replicate inside bacterial cells independently of the bacterial chromosome, and express their own genes.

So how did they produce the different mutant versions of the ferritin gene? This brings us to one of the coolest parts of the study; they did it via error-prone PCR. They used a faulty PCR process, taking advantage of an error-prone DNA polymerase, which introduced random changes into the ferritin gene that was being copied. This allowed them to produce hundreds of randomly mutated versions of the ferritin gene. Each mutant ferritin gene was inserted into a plasmid as stated above. The most magnetic of these mutants were then isolated from the total mutant population by magnetic column retention characterization: the cells suspended in buffers were passed through a magnetic column of high magnetic field gradient placed between two permanent magnets; the most magnetic cells remained bound to the walls of the column while the cells that failed to bind were flushed out by washing with more buffer solution. 

The isolated magnetic cells were grown and then passed through the magnetic column again to select for the more magnetic cells. This was repeated ten times as part of the process of directed evolution: providing selection pressure on the cells to facilitate evolution of the greater expression of a certain trait, in this case, magnetism. After thus evolving highly magnetic cells (presumably containing mutations in the ferritin gene that enhanced magnetism), the mutant cells were plated, and several of them were sequenced thoroughly – this can be defined as the process of mutant screening in the concept of forward genetics. They were thus able to figure out the exact mutations required in ferritin to increase the magnetism of the assembled iron oxide core. To add to this understanding, they then applied the principles of reverse genetics, in that they introduced each of the mutant genes into the initial wild-type ferritin gene on a plasmid in order to verify whether the change in iron-sequestration was solely due to these mutations.

In total, 38 mutants were tested, and the results revealed that the majority of these significantly increased the magnetism of the cells compared to wild-type ferritin. Also, the most magnetic mutant is seen to display double point mutations which change the amino acid configuration of the protein constituting the B-type channel, through which iron molecules enter the ferritin molecule. The mutated B-type channel appears to be changed in shape to facilitate increased iron influx into the ferritin core region. Via SDS-PAGE gel, it was observed that the mutant ferritin genes had lower protein expression compared to the wild-type ferritin gene, confirming thus that the increased iron-sequestration was due to increased magnetism and not due to more protein being present to work. Interestingly, the mutants were also observed to exhibit the startling ability to sequester other metal, namely zinc, cobalt, and nickel.

To say that that this experiment was beyond thorough in verifying the genotype-phenotype link would be an understatement. Aside from checking magnetic retention, the researchers adopted a handful of other methods to make confirmatory observations. For instance, MRI and fancy techniques such as SQUID Magnetometry (basically a very sensitive device for detecting and measuring magnetic fields) were used, again, to check degree of magnetism, all of which showed favorable results. The mutant cells were also placed in welled-plates directly above cylindrical magnets and were observed to take the cross-sectional shape of the magnets below them in alignment to the magnetic field. Overjoyed, the scientists remembered to take snaps of this plate – a lovely picture in the memory (and as evidence) of an undoubted success.

The discoveries from this elaborate study spell out the opposite of Magneto’s aspirations – the benefiting of mankind using mutants as pawns. Indeed, the evolved magnetic protein, with a little help from biosynthetics, could be applied to countless bio-applications. A simple example could be the utilization of genetically modified cells which attract and store toxic or valuable metals for bioremediation (neutralization of harmful waste from contaminated sites using microorganisms) or mining. The magnetic ferritin gene could also be used as a genetically-encoded reporter gene – i.e. as a genetic tag which can be made to be expressed with the gene being studied. The greater the expression of the gene, the higher the amount of magnetism of the cell due to the co-expression of the mutant ferritin. In addition, ferritin mutants could function as non-invasive reporters of biological signals from engineered cells, for instance, immune cells targeted towards a cancerous tumor (you could trace them by their magnetic fields).

Given that the natural size of the ferritin cage – a miniscule 8 nm in core diameter- is too small to create sufficient magnetic moment to acquire optimum sensitivity and reliability, all of this is considerably ambitious. However, like with DUF892, which is a similar protein to ferritin that they discovered during this study, there lies promise in other similar proteins in discovering a solution to overcome this shortcoming. One slight limitation of the original paper is that they do not explain why we cannot just use the magnetic properties of magnetotactic bacteria (which use the earth’s magnetic field to direct themselves) for our purposes, despite mentioning that they assemble more crystalline and magnetic iron oxide. One possible reason for that is that the magnetic properties of magnetotactic bacteria are conferred by membrane-bound organelles called magnetosomes, which contain several transmembrane proteins. Ferritin is therefore a much simpler system to work with, or insert into different bacteria. Besides, they were also looking at what properties of an individual iron-binding protein can be modified to enhance its magnetism.

There’s a saying that power is neither good nor evil- only the nature of its wielder decides that. While Magneto remains a fictitious threat to our comic-book heroes as well as mankind itself, the tiny mutants possessing his powers in the real world give hope for amazing applications in biology. So let’s end this piece wishing prosperity to the more innocent, bacterial versions of the antihero – may they flourish in future research and take us further.

Maliha is a weirdo who somehow believes she's from a different planet. But she likes Earth just fine, and is fascinated by the science and beauty of life and has made it her purpose to explore it. Besides this, her most burning desires include becoming a synthetic biologist/ genetic engineer and running away with a heavy metal band.