Buddy Breathing

We all need to breathe. This is true for us humans, and it’s true for most microbes. However, many microbes have different approaches to breathing. A new discovery from Derek Lovley at University of Massachusetts reveals surprising flexibility in microbial respiration—and how a couple of bacteria can cooperate and evolve to meet a challenge to their survival.

When we breathe, we’re consuming oxygen; we need this oxygen to oxidize our food, which is mostly carbon and hydrogen (think “carbohydrates”). We often think of oxidation as simply “reaction with oxygen”. After all, we are taking the carbon and hydrogen we eat and making it into carbon dioxide and di-hydrogen oxide (or water, for the less nerdy). However, there is a more precise definition of oxidation that appeals to chemists and microbiologists. Oxidation is the loss of electrons by an atom; since the electrons have to go somewhere, they are accepted by another atom, which we say is getting reduced.

When we respire (which is why we breathe), we are taking electrons away from the carbon and hydrogen in our food, and giving those electrons to oxygen. We get energy from this process, which we use to make the ATP that powers the cell. But Bacteria and Archaea have figured out how to take electrons from carbon and hydrogen and give them to atoms other than oxygen: sulfur, iron, arsenic, and all manner of other atoms. They don’t necessarily get as much energy out of these processes, but it’s better than dying if no oxygen is available.

One example of this type of anaerobic (no-oxygen) respiration is seen in the Bacterium Geobacter metalloreducens, which, as its name suggests, reduces metals. When it respires, it “eats” ethanol, taking away its electrons, and “breathes” metals such as iron, giving them electrons and producing rust. Another example of this type of respiration is found in the related bacterium Geobacter sulfurreducens. It can’t eat ethanol, but it can eat hydrogen. It can’t “breathe” oxygen or iron, but it can give electrons to sulfur (as the name suggests) or the compound fumarate.

Whether or not oxygen is used for respiration, the basic outline of the process is the same: electrons are plucked off of the food, and go through a series of intermediate carrier molecules before they end up at the terminal electron acceptor. The whole process takes place in the cell membrane: the electrons on their carriers never leave the cell until they are handed to the terminal electron acceptor. As the electron is passed from carrier to carrier, it releases energy, which is used to make ATP.

The outline of this process is the same for us, and for Geobacter metalloreducens and for its cousin sulfurreducens. However, metalloreducens and sulfurreducens are specialized for particular combinations of food and electron acceptor: metalloreducens likes ethanol and iron, while sulfurreducens likes hydrogen and fumarate.

But what happens if you put these two organisms in an environment with food for only one, and an electron acceptor for only the other—ethanol and fumarate? It would seem that metalloreducens could take electrons from ethanol, but not get rid of them, while sulfurreducens could give electrons to fumarate—if only it could obtain some electrons from somewhere!

Fortunately, these two different types of cells can cooperate. Geobacter metalloreducens can get rid of electrons from ethanol—it just won’t get much energy out of the deal. Water, it turns out, is mainly H₂O, but always has some OH- and some H+; metalloreducens can give those electrons from oxidizing ethanol to H+, making hydrogen gas. This works out nicely, since sulfurreducens can “eat” the hydrogen, and give the electrons to fumarate. So, even though each member of the partnership only gets half as much energy, it’s better than getting no energy at all.

This type of collaboration is called “syntrophy,” and it’s actually fairly common in the microbial world. One organism respires, producing hydrogen; the next organism respires, “eating” that hydrogen. Each organism has a complete respiratory system, but the two respiratory systems are dependent upon each other. Syntrophy occurs in your own personal digestive system, in which one organism makes hydrogen, giving electrons to another organism, which uses them to make the methane that makes our farts flammable. Up till now, when syntrophy has been studied, it has always involved hydrogen as the electron carrier. However, Lovley’s group introduced an interesting complication to this process.

Lovley’s group mutated sulfurreducens so that it absolutely can’t eat hydrogen. They then started the experiment again, giving a mixture of metalloreducens and sulfurreducens nothing but ethanol to eat and nothing but fumarate to breathe. Given that the options for these populations of cells were (a) evolve or (b) die, it’s unsurprising that option (a) was chosen.

What was surprising was the way that evolution worked. Rather than jury-rigging a way for sulfurreducens to eat hydrogen, evolution chose a method that involved the direct transfer of electrons from metalloreducens to sulfurreducens. I mentioned that when a cell respires, the electron gets shuttled from one carrier molecule to another as it moves from the “food” to the electron acceptor. These carrier molecules stay in the cell membrane, and respiration happens strictly within the confines of the cell.

What Lovley found was a mutant form of sulfurreducens that produced electron carriers that could be exported to cells of metalloreducens, pick up electrons, and then shuttle back to sulfurreducens. The sulfurreducens cell had to make filaments for these carriers to travel along, and had to grow next to metalloreducens—but one part of respiration was happening in one cell, and the other part of respiration was happening in another cell. This is syntrophy, but without using hydrogen to transfer the electrons. Electrons were flowing directly from one cell to the other.

Being thorough, Lovley’s group made sure that they were seeing electrons moving directly between the two cells. They checked to see if hydrogen was being exchanged—it wasn’t. They found the genes in sulfurreducens that had mutated to allow this to happen, and deleted them—and growth stopped. They looked for electrical current between cells—and found up to four microamperes of electron flow. They looked at the cells using various forms of microscopy, and found that they grow in close contact…

(this is a clump of cells, with metalloreducens stained green and sulfurreducens stained red. They can't grow separately, like the cells in a normal syntrophic relationship.)

…and they used the much higher magnification of electron microscopy to show that sulfurreducens made a web of filaments that connected it to metalloreducens cells, and that electron carriers (visible as dark spots) could move along those filaments:

This has never been seen before—respiration has always been contained within one cell. But now, we know that the process of respiration can be completely shared between two cells, a sort of bacterial “buddy breathing.” One cell oxidizes food (metalloreducens taking electrons from ethanol) and the other reduces an electron acceptor (sulfurreducens giving electrons to fumarate). This kind of metabolic sharing allows both cells to survive in an environment that could not sustain each individual, and so gives life the ability to exploit ever more of the earth’s environment. It’s also a nice example of life’s seemingly endless creativity—what Daniel Koshland calls “improvisation”—using mutation and selection to try out novel solutions to novel problems.

Zarath M. Summers, Heather E. Fogarty, Ching Leang, Ashley E. Franks, Nikhil S. Malvankar, and Derek R. Lovley (2010). Direct Exchange of Electrons Within Aggregates of an Evolved Syntrophic Coculture of Anaerobic Bacteria. Science 330: 1413-1415.

Koshland, Daniel (2002). The Seven Pillars of Life. Science 295:2215-6.