Biology, Chemistry, Physics | UNDERSTAND

Titanic and the iron-eating A species of bacterium discovered on the decaying wreck of the is providing new insights into how to protect living cells from damage.

By Giuseppe Zaccai

The wreck of Titanic lies on the ocean floor, nearly 4 km located in the deep sea. However, these organisms have now below the surface of the dark, salty water of the North become the focus of scientific research for another reason: to Atlantic. Divers on expeditions to the site have noticed find out how they manage to thrive in conditions with high ‘rusticles’ – icicle-like growths rich in rust – covering parts and varying salt concentrations. of the damaged and corroded hull. In 2010, scientists investigating rusticles taken from the Titanic site discovered a Salts and the living cell new species of bacteria within these growthsw1. They named The term halo- means ‘salt’ in Greek. Bacteria of the genus the new species titanicae. Halomonas live in salty environments, such as sea water The scientists have also discovered that Halomonas or salt marshes, where salt concentrations vary over a wide titanicae bacteria are involved in the rusting process and are range. All members of the genus are strongly ‘halotolerant’: accelerating the wreck’s decay. The decay is happening at they have evolved ways of coping with wildly fluctuating salt quite a rapid rate: estimates suggest that the wreck may have concentrations outside the cell, surviving over a huge range disappeared completely by 2030. These bacteria could thus of salinity from 0.5% (w/w) sodium chloride right up to 25%. pose a significant threat to oil rigs and other iron structures This may seem like a trivial issue, but it is not.

The RMS Titanic at the docks at Southampton, UK in April 1912 Wikimedia Commons (public domain)

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Microbiology Chemistry Image courtesy of Lori Johnston, RMS Titanic Expedition 2003, NOAA-OE Biochemistry Archaeology Ecology Ages 16–19

The is very well known, so this article can be used to excite students about science. It can also act as an incentive A view of the bathtub in the captain of the Titanic’s bathroom. Rusticles can be seen growing over most to delve deeper into of the pipes and fixtures in the room. the world of bacteria and their usefulness to Cells are protected from the outside from the more dilute side to the more human beings – despite world by very thin membranes that concentrated side. their normally negative associations. control traffic in and out of the Cell membranes cannot cope with The article would cell, creating and supporting huge osmotic pressure differences. Red be useful in biology differences in composition between blood cells will burst when placed in teaching to explore how the inside and outside of the cell. For pure water, illustrating the effect of prokaryotic organisms example, cells maintain an electrical osmotic pressure – and the fragility can survive in extreme potential of about 100 millivolts across of cell membranes. Clearly, to keep environments, and how the membrane, which is typically just their membranes intact, Halomonas substances are transferred 10 nanometres thick. For all types of bacteria must be able to regulate across the cell membrane. the concentration of solutes in the It could also be used cytoplasm in response to changing in chemistry teaching external salt concentrations, without All Halomonas about hydrogen bonds disrupting the biochemistry within the “ REVIEW and the interactions bacteria have evolved cell (see Zaccai, 2009). How between water and do they do this? ways of coping with other molecules. wildly fluctuating salt Halomonas bacteria produce large Finally, recycling using amounts of a substance called ectoine Halomonas titanicae concentrations outside (see figure 1). To counterbalance the is relevant to ecology the cell. external salt concentrations, they and the study of the ” adjust the concentration of this soluble energy balance within compound within their cells. This keeps ecosystems. the cell’s fluids in osmotic balance cell, sodium chloride (NaCl) is the with the outside, protecting the cell Alina Giantsiou-Kyriakou, dominant salt outside the cell (for against shrivelling up or bursting even biology teacher, example, in blood serum), whereas in extreme conditions. Remarkably, Livadia High School, potassium chloride (KCl) is dominant ectoine appears to act differently Cyprus inside the cell. Yet, while the salt types from most salts and solutes, which are different, the total salt concentration can interfere with the role of water on either side of the cell membrane in metabolic processes. Ectoine is a should match exactly. The reason for ‘compatible solute’, and so preserves this is osmosis: if two solutions of the normal biochemistry within the cell. different concentration are separated It also increases the stability of proteins by a membrane that is permeable to and membranes. So how does this water but not to solutes, water will flow unusual solute achieve these results?

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Ectoine and the membrane surfaces (Zaccai et al., The attraction between water molecules hydrogen-bond network 2016). Normally, water molecules is also the basis of the ‘hydrophobic interact with each other through a effect’, which makes oil and water Clues as to exactly how ectoine works network of hydrogen bonds. The atoms separate. The hydrophobic effect is on the molecular level have recently in a water molecule interact with important in many biological processes, been discovered using the advanced neighbouring molecules, with each including the three-dimensional folding experimental technique of neutron oxygen atom acting as a receiver of two of protein molecules and the formation scattering (see text box). Carried out hydrogen bonds and each hydrogen of membranes. Hydrogen bond by an international collaborationw2 atom as a donor of one bond. This dynamics are thus an essential factor in involving the Institut Laue-Langevin results in a highly dynamic network how living cells are organised. (ILL)w3, the experiments showed how of intramolecular bonding, with the ectoine influences the layer of water molecules changing partners a billion around protein molecules and on (109) times a second. “ Hydrogen-bond dynamics are an Neutron scattering essential factor in Neutron scattering is a powerful method for studying the structure of how living cells are materials, including water and its interactions with other compounds. organised. The nucleus of a hydrogen atom is a single proton. Neutrons and protons ” are very similar particles (apart from their electric charge), and neutrons strongly scatter off protons, rather like snooker balls bouncing off each other. The neutron scattering pattern provides information on the Other substances in the water, location of oxygen and hydrogen atoms in nearby water molecules, and such as salts, can interfere with this thus on the dynamic network of hydrogen bonds that links them. organisation. But, as the neutron Another useful trick is used in neutron scattering. The nuclei of experiments have revealed, ectoine deuterium atoms (an isotope of hydrogen with one proton and seems to enhance hydrogen-bond one neutron in its nucleus) scatter neutrons in a very different way dynamics – at least in part – rather than from normal hydrogen nuclei. Replacing the hydrogen in a specific hindering it. This molecule contains a compound (ectoine, for example) with deuterium makes it possible positively charged group (shown in blue to see the contribution of only this compound to the scattering signal, in figure 1) and a negatively charged distinct from that of the surrounding water molecules. This in turn helps group (shown in red). These charged to reveal the details of their interaction. groups interact with water molecules, Neutron scattering with deuterium labelling has provided a rich crop making the hydrogen bond network of results in structural biology. These include insights into the complex nearby a little less dynamic – but also interactions between protein molecules, and those between proteins and creating a more dynamic network of nucleic acids (DNA, RNA). water molecules further away.

CH3 N

NH

OH

H ©ILL/L. Thion

BACKGROUND 0

Figure 1: The chemical structure of ectoine. In water, The D22 instrument at ILL, which revealed how water and ectoine interact with the red group becomes negatively charged (by losing + protein surfaces. The door of the apparatus has been opened to show a view of the one H ion) and the blue group becomes positively + neutron detector. charged (by gaining one H ion). The charged groups then form hydrogen bonds with adjacent water molecules.

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Outside the bacterial context, ectoine References w3 The Institut Laue-Langevin (ILL) is an international research centre at the leading is proving to be a useful ingredient Zaccai G (2009) The intracellular environment: edge of neutron science and technology, in cosmetics and clinical treatments, not so muddy waters. Science in School 13: based in Grenoble, France. See: www.ill.eu because it can reduce inflammation in 19-23. www.scienceinschool.org/2009/ issue13/water mammalian cells. The likely reason for Resource this is that by stabilising proteins and Zaccai G et al. (2016) Neutrons describe ectoine effects on water H-bonding and Find out more about microbes that live on membranes, ectoine protects human hydration around a soluble protein and a shipwrecks and the role of ectoine in an cells from damage, thus reducing cell membrane. Scientific Reports 6: 31434. accessible article on the BBC website. See: inflammation and providing remedies doi:10.1038/srep31434 www.bbc.com/earth/story/20170310-the- for conditions such as allergies, eczema, wreck-of-the-titanic-is-being-eaten-and- may-soon-vanish and cough and cold symptoms. Web references ______w1 Halomonas titanicae bacteria were discovered Return to the Titanic by scientists from universities in Seville, Spain, and Toronto and Halifax, Canada. Joe (Giuseppe) Zaccai is Emeritus Although the neutron experiments Read a scientific account of the discovery on Director of Research at the National have helped scientists to understand the UNESCO website. See: www.unesco.org/ new/fileadmin/MULTIMEDIA/HQ/CLT/pdf/ Centre for Scientific Research (Centre how Halomonas bacteria thrive in the Henrietta_Mann_Paper.pdf National de la Recherche Scientifique, potentially hostile environment around w2 The collaborating organisations included the CNRS) in France. He spent his research the sunken Titanic, the exact role of Institut Laue-Langevin (ILL) and Institut de career based at ILL, where he developed Halomonas titanicae in rust formation Biologie Structurale in Grenoble, France, and neutron scattering methods and applied remains unclear. But there is some the Max Planck Institute of Biochemistry them to the structural biology of good news here: these iron-eating and biotechnology company Bitop in Munich, adaptation to extreme environments. bacteria could have a role in future Germany. See the online supplementary information section in Zaccai et al. (2016) for waste management, speeding up the more information. decomposition of metallic litter – aside from historic wrecks – on the ocean floor.

Deepsea Delta oil drilling rig in the North Sea. Like the wreck of the Titanic, oil rigs are at risk of damage from rust-eating bacteria. Erik Christensen/Wikimedia Commons

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