How'd he DO that? Real Chemical Magicians

Enzymes 101: Peaking into the black box

            The term “enzyme” has a pretty specific meaning to people who define themselves by such terms as “enzymologists” (go figure). To most people, however, it’s a less well-understood term. For many of the folks I know, the term “enzyme” can be virtually interchangeable with “protein,” or perhaps only recognized in the context of specific and dubious dietary supplements. For the purposes of this and further discussions, I’d like to talk about enzymes as proteins or nucleic acid that act as a catalyst to speed along chemical reactions. That’s what enzymes are and what they do, but how on earth do they do it? In this post we’ll use broad strokes to discuss some of the general strategies enzymes use to put eons of biochemical evolution to work.

            It can be a bit hard to really appreciate, and I mean fully appreciate, what enzymes do for us, and how difficult this task is chemically. When I think of the power of enzyme chemistry it always brings me back to early organic chemistry classes. As a bright-eyed, eager young undergrad I dutifully learned the rules of traditional organic chemistry: what bonds can be broken by what chemicals under what impossibly harsh boiling acidic conditions. We learned that by the natural laws of energy, matter, and all that is chemically holy there are just certain reactions that wouldn’t happen at room temperature during the length of an average PhD. How then, I would ask, could these types of reactions be magically popping up in textbooks looking perfectly and innocently plausible? The professor’s answer? Enzymes.

            Upon receiving such a disyllabic and seemingly self-evident answer I’m sure that I nodded my head knowingly, pretending that I fully understood how the biochemical black box could perform chemical black magic. Since then I’ve made a few strides toward actually understanding the mechanisms behind these baffling enzymatic chemoacrobatics, and continue to be amazed by the underlying simplicity and elegance in complicated enzyme systems. As an introduction to the broad concepts of enzyme mechanisms I’ll talk about the very basic catalytic strategies they use, and leave more detailed mechanisms for a later date. So let’s talk about shape, strain, orientation sequestration, and transformation. Lets talk about enzymes.

            One of the ways that enzymes are able to shift chemical reaction rates into high gear is simply by bringing the necessary components, or substrates, together. Understandably, this seems trivial on the surface but practically it is both very important and very effective. Being macromolecules (proteins or nucleic acid), enzymes have lots of different chemical groups: hydroxyl groups, aromatic groups, amine groups, etc. These have been positioned during evolution of specific enzymes to be in just the right places to interact very specifically with their substrate molecules. The “lock and key” analogy is used a lot in explaining these interactions, and is as apt an analogy as any. Basically the enzyme is able to bind very specific compounds, while completely ignoring others. By doing this for more than one substrate, they are able to bring two or more reacting molecules into very close proximity and hold them there, allowing them to interact much more efficiently. Additionally, enzymes don’t bind their substrates in just any old way, but in a precise orientation, effectively lining up slot A with tab B. This very simple enzyme function is kind of like a good match maker, taking the chance out of two compatible things coming together and setting the mood just right for the chemistry to happen.

            Another power that enzymes have is a consequence of their size; enzymes are huge compared to the small molecules involved in many of the reactions they catalyze. This means that they can completely surround these small molecules, and in so doing can control the immediate environment in which the reaction is taking place. Practically this can mean separating the substrates from interfering water molecules or creating pockets of positive or negative charge in specific places. This gives the substrate molecule(s) a custom-built productivity space where all of the elements are designed to help the chemistry along and exclude interfering substances.

            The general mechanisms we’ve talked about so far involve concepts that I like to think of in terms of “tweaking the chemical circumstances,” but enzymes can play a much more “active” role in chemical reactions as well. One way that enzymes can shove a reaction in the right direction is an offshoot of the “lock and key” tight interaction concept mentioned above. Some enzymes bind their substrates in such a way that they actually bend, twist, or otherwise strain their shape. This changes the energy state of the molecule and helps to push the chemical reaction to its conclusion. Alternatively, the enzymes themselves can actually react chemically with their substrates, forming a covalent bond between enzyme and substrate. This bond is broken by the end of the reaction cycle to release the product and regenerate the enzyme. This latter mechanism is often referred to as providing an alternate path for the chemical reaction to go through, effectively using the substrate-enzyme reaction as a handy detour to avoid more difficult chemistry.

            The concepts of catalytic mechanism explored above are incredibly general, and the specifics that happen in each individual system can be complex enough to fill the contents of a research career (or several). With all of the complicated and truly challenging chemistry that’s going on in our bodies every day, however, I think it’s inspiring that such simple and intuitive concepts are at the root of such complex mechanisms. It’s amazing what we can understand about our own chemistry if we choose to start looking inside the black box.

DNA Repair just doesn’t give me the same Buzz

Ask any graduate student and they might tell you caffeine is a lifeline. Ask a health enthusiast and they might tell you it’s a poison. Ask a physician and between sips they might advise you that it’s fine in moderation. If you decide to ask the researchers behind a recent paper in the Proceedings of the National Academy of Sciences, however, they’ll let you know that caffeine can help prevent skin cancer. As a bonus, they’ll even tell you why. In my post last week I started what I intend to be an ongoing discussion on enzymes: what they are, what they do, and how they affect every facet of life. The last post was a (very) general introduction. Some posts will dig into the nuts and bolts of the chemical reactions enzymes help along and how the heck they do it. Some posts, like this one, will discuss the broader roles of enzymes in biology. So grab your coffee cup and sunscreen (just in case) as we talk about some counterintuitive caffeine consequences.

            Before we can talk about what role enzymes play in cancer, we need a very brief description of how cancer works. Cancer occurs in pretty much all life forms that exist, like us, as groups of cells. This is in contrast to single-celled organisms like bacteria and yeasts. Cancer is basically uncontrolled growth of specific cells. All the cells in our body usually divide to form new cells, thus growing the tissue, at specific rates and under specific circumstances. Cancer happens when the cells divide very rapidly without obeying the rules, so to speak, of when they are supposed to replicate. One of the reasons that normal cell division is so regulated is to make sure that the new cells coming out of cell division are healthy and have accurate copies of the parent-cell DNA. When replication is happening too quickly, there is no time for the cell “quality control” mechanisms to check that everything is honky dory, and the result can be new cells with mistakes in the genetic code. These aberrant cells not only have functional problems due to the mutations, but will also go on to divide rapidly, causing a cascade of rapidly dividing, unhealthy cells that form the tumors associated with cancer.  So what is the trigger for this cascade? What causes that initial cell to start dividing too fast? As I mentioned, normal cell division is closely regulated, and if something causes a problem in one of the tools the cell uses to regulate division, the regulation system can go out the window. The genes coding for these regulatory tools are often called oncogenes (basically “cancer genes”) as mutations in these genes are likely to cause cancer.

            There are many things coming at us every day that can cause DNA damage. These range from UV-rays to charbroiled steak to chemicals we make inside our own cells or mistakes by our cellular DNA-manipulation machinery. In fact, lots of DNA damage is done every day inside each one of us, so why are we still up and walking around? Enter the enzyme. Not one enzyme, in fact, but an arsenal of enzymes, each with a specific job to do in DNA-maintenance. In thinking of enzymes in your body, you can think of each one having a very specific skill, like trades-people working to build a house. The plumber doesn’t put in the electrical work and only the floor guy puts in the tile. With enzymes it goes even further, so that in laying floor tiles you’d have one guy to lay the grout, one guy to pick up the tile, another to position it, another to press it down, another to wipe it clean, etc.  In talking of DNA, there is a set of enzymes for making the DNA, specific sets of enzymes to repair specific types of DNA damage, and specific sets of enzymes to detect specific types of damage at specific times and signal to the DNA-repair enzymes to get to work.

            Before we get too jittery, lets talk about how caffeine affects this process. We all have an enzyme called ATR that is involved in a couple things we’ve talked about. This enzyme is a kinase, meaning that it catalyzes transfer of a phosphate group from one molecule onto another. This might seem a bit inconsequential in the context of something as huge as cancer. This one transfer reaction, however, is a recognizable signal in the cell and is passed along and amplified. Eventually it triggers the action of enzymes tasked with repairing certain types of DNA damage, including that caused by UV-rays. The enzyme ATR also happens to be part of the division regulation “tools” that we talked about. It’s a kinase that performs its role as part of a cell division checkpoint, a time when activities in the cell determine if it will go on to divide, or kill itself in a process called apoptosis.

Caffeine binds to ATR and stops it from doing its job. This means that when some kinds of DNA-damage is detected, ATR does nothing (instead of transferring that all-important phosphate), the DNA is not repaired, and instead of replicating, the cell dies. Wait a second, this sounds like a bad thing; how does this prevent cancer? The problem with DNA repair enzymes is that for certain types of DNA damage, there is no way for them to ensure that the DNA is put back together exactly like it was before the damage. Sometimes these enzymes can only physically fix the break and hope that the sequence is repaired by luck, or that it was in a spot that didn’t matter much anyway. If this type of repair happens in an unlucky spot, like an oncogene, the repair makes DNA that looks physically okay, but the resultant mutation can have cancerous consequences. In these cases, NOT repairing the DNA effectively causes cellular suicide before the very first cancer cell can form.

Enzymes have a role in everything our bodies do, from detecting signals and passing messages, to constructing and repairing cellular components. Everything is controlled in a delicate balance, and often this control is itself achieved by enzymes. As this example illustrates, turning an enzyme “off” is an important component of cellular control mechanisms. Although our bodies have many built-in off switches, outside chemicals can also interact with our enzymes with ultimate results that can be difficult to predict. So next time you’re chowing down you can look at your food and ask, “Hey, what enzyme are you hooking up with?”

It’s a Molecule, it’s a Chemist, it’s Super Enzyme!

Photo credit whatsthesmatter@blogspot.com

Enzymes 101: The Introduction    

       

It’s amazing how many things our bodies can accomplish without us ever realizing that it’s happening. This is especially amazing when we consider that many of these tasks are absolutely vital to keeping us alive and conscious enough to appreciate it. How many times a day do you notice that you’re blinking? How often do you register your heart beating? For many, the answer is probably rarely (although probably now that you’re thinking about it). There are thousand of actions going on in your body, however, that you will never notice directly, no matter how hard you concentrate on them. These are the chemical reactions that make the difference between semi-organized piles of organic mush, and the living, breathing, navel-gazing individuals we all are. So if you’re happy and you know it, thank your chemistry; without it I can guarantee neither condition would be true.

Some of the chemistry that occurs in our bodies is as simple and spontaneous as the old vinegar and baking soda volcano trick. Any reaction that explodes into reality in a fourth-grade classroom is hardly an energetic challenge. Other reactions, however, are a lot tougher to coax along. Even when some important reactions do occur spontaneously (that is, without other factors interfering in the process) the speed of these chemical reactions, or reaction rate, might be so slow that it wouldn’t be likely to happen over the course of one’s lifetime, much less the multiple times a second required for it to be biologically useful. With all of this difficult chemistry so necessary to stay alive, how do we accomplish these constant chemical feats? Enter the enzyme.

Before discussing the amazing power of enzymes as tools or, as I like to think of them, master chemists, we need to go over a few defining concepts. Enzymes are catalysts. What this means is that although they help to chemically transform something into something else, they emerge from that reaction just the same as when they went in. This is like a baker: without him there is a very VERY small chance that all of the ingredients would jump into the oven together and become a cake, but while they come out transformed into a different and delicious form, he stays the same and is ready to bake another cake. What this means for your body is that unlike sugars, oxygen, and numberless other chemicals that your body has to constantly take in to supply the “ingredients” for your life-chemistry, a single enzyme can contribute over and over to the same chemical reaction without having to be replaced.

Now we have a bit of an idea of what an enzyme’s role in a chemical reaction is, but what is the enzyme itself? Enzymes are macromolecules made up of protein or nucleic acid (or both). A macromolecule is basically what it sounds like: a big molecule. Proteins and nucleic acids are big molecules made up of chains of smaller units. These chains then fold up into a shape that gives the enzyme its function. We’ll talk more about the specific forms and families of enzymes in future articles.

The more you learn about nature, the more you learn what a spectacular problem solving force evolution can be. When it comes to chemistry, the problem-solving power of enzymes is more astounding than the most outrageous magic any fiction writer could ever come up with. So stayed tuned for future posts when I’ll explore the magic mechanisms, resplendent reactions, and elegant evolution of nature’s best chemists!

Is that a Fruitloop in your Backpack?

As any six-year-old can tell you, toucans are named Sam, speak English, and push mini fruit doughnuts like they’re going out of style. As any ornithologist can tell you, toucans are jungle birds playing a vital role in helping spread the seeds in their favourite fruits far from the parent plant. As any geocacher can tell you a good GPS unit can save you a lot of crawling on your belly through the underbrush. Recently a group of scientists combined the latter two nuggets of wisdom to develop tools for studying avian roles in forest dynamics. Turns out you can learn a lot more from Toucans than what passes as part of a complete breakfast.

            Like all other forms of life, plants employ a wide variety of strategies to ensure the success of the next generation and the propagation of their own genes. Whereas human parents tend to perform two am feedings and invest in high quality car seats, members of the plant community are more limited in their ability to nurture their descendants. Indeed, the extent of plant “parenting” is often limited to giving their seeds the best possible chance of settling on a great growing location. Unlike human infants, the best place for young plants tends not to be under the watchful leaf of their parent plant but much further afield. There are several reasons that a little more separation is often the right answer for a seedling plant. Firstly, competition for resources like light, water, and soil nutrients is often higher around the base of their parent plant. There are also plant parasites like insects and fungi that tend to congregate around the older plants.

            In order to give young plants the autonomy so desperately desired by myriad human teenagers, plants produce seeds with built-in travel mechanisms. Some seeds have sails or propellers for dispersal by wind, some float for water dispersal, and still more have hooks to catch on the fur of animals or the socks of hikers and be carried away. By far the most delicious solution, however, is encasing seeds within fruits. In this case animals are able to eat the fruit and unwittingly scatter the tougher seeds as they travel, often with a batch of home-made fertilizer. This is the strategy used by the tropical Virola nobilis tree, the fruits of which are frequent breakfast choices for toucans.

            While seed dispersal is an important key to understanding forest ecosystems, studying how animals and birds disperse seeds by tracing either party can be difficult or even impossible. To get around the technical limitations inherent in clumsy, flightless humans trying to follow swift, winged toucans, a group of scientists employed some pretty slick technology. This group attached mini-backpacks equipped with GPS and 3D accelerometer units to wild toucans to study the feeding habits and movements of the toucans. The clumsy humans, instead of swinging through the trees, used the accelerometer readings to determine when the birds were eating, used the GPS readings to determine when and where they were traveling, and used zoo-dwelling toucans snacking on the same food to determine how long after eating they generally regurgitated the seeds. Using these methods, researchers can study how bird populations contribute to the overall makeup of the forest.

            Ecosystems, like forests, are truly a community effort. Every plant, animal, bacterium, fungus, and forgotten grad student plays a specific role in the maintenance of that ecosystem and the survival of all the other species. It’s no breakfast cereal cash cow, but through seed dispersal, real toucans are a vital piece of the forest ecosystem puzzle.

Dude looks like a Lady

If anyone can tell us how far we’ve come from Victorian views of sexuality, it’s the makers of women’s undergarments. As we’ve shed our voluminous petticoats and embraced the “man bag,” we’ve also started to accept that human sexuality has a few more shades of gray than may have been acknowledged in days of yore. While it’s not much of a stretch to use clothing to blur gender boundaries, you might be surprised to learn just how delicate your physical gender can be. When it comes to gonads, it really can be the cut of your genes that matters.

            And now for the basic biology version of the hated “birds and the bees” discussion: Mom provides the egg (one set of genes), and Dad provides sperm (a second set of genes). When the two meet up, under circumstances best described by parents the world over, they fuse and the two sets of genes form the necessary DNA to make a whole little person. The genes in question are in bundles called chromosomes (23 pairs in human cells), and the sex of the bouncing baby is determined by whether they carry the X or Y flavour of the sex chromosome. As all women have two X chromosomes and men have one X and one Y, the sex of the baby is dependant on whether the lucky egg-meeting sperm is of the X or Y persuasion. So the gender of the baby is determined, forever and always, by the sperm that wins the race, right? Normally, yes, but biology also has some tricks up its sleeve.

            The reason that the X and Y chromosomes are so all-important in gender decisions, beyond their nifty alphabetical designations, is the genes they carry. Some of these encode for proteins that produce sex hormones. Hormones are small molecules in your body, and the ones from the ovaries and testes are responsible for all the differences between male and female bodies. Incredibly, every function, feature, and fashion sense that defines our genders boils down to the initial decision for a single cell type to become either Sertoli cells (which are part of the testicle) or ovarian granulosa cells. These cells later play a key role in production of gender-appropriate hormones, resulting in development of everything from genitalia in the womb to scruffy adolescent mustaches.

            If the difference between developing Sertoli cells and granulosa cells is the rather important one of gender, surely this process occurs early and is promply cast in stone, right? Actually, like so many things in our body, the cellular decision to develop into one cell type or the other is controlled by a balance of inputs from two opposing “male” and “female” signaling pathways. Even more surprisingly, regardless of the identity of that initial sperm, if key components of one pathway are disrupted, the opposite gender will win out. That is to say that even if someone is genetically male (XY), if there is something wrong with this signaling pathway from the Y chromosome, they will develop like a female, and vise versa. If that gender jack-in-the-box doesn’t knock your unisex socks off, consider that new research indicates this can happen at any point in life. A recent paper describes how disrupting the “male” pathway in adult mouse testes resulted in these organs actually turning into ovaries.

            These experiments don’t offer practical applications, nor is this at all likely to happen spontaneously (don’t worry guys, you can stop clutching your pants in terror). They do, however, give us fascinating insight into what our genders really are and where they come from. Boy meets Girl, and Sperm meets Egg, but the new life emerging from those unions always looks just like its real Dad: biochemistry.

Glowing Green Mushrooms Seem Pretty Magical to Me…

            If you ask me (or any mycologist) the term “magic mushrooms” isn’t nearly specific enough to describe the psychedelic variety usually implied. The truth of the matter is that lots of mushrooms are pretty darn magical. Aside from providing a tasty side dish, these fabulous fungi produce an astounding array of chemical compounds with powerful effects. From delectable truffles, to potent poisons, many mushrooms have a profound ability to spice up an otherwise bland afternoon. One of the most amazing fungal feats, however, is best appreciated not by eating, but by looking. That green glow isn’t in your head, it’s the forest night-light: fungal bioluminescence!

            Before delving into their Lite-Bright-like capabilities, we should answer the question: what exactly is a mushroom? When it comes to the world of fungi, our umbrella-shaped friends are literally just the tip of the iceberg. The mushrooms we see growing above ground (and appearing on our dinner plates) are only one part of a larger fungal organism. The Fungi are a very diverse group of organisms and have many shapes and forms. Most fungi, however, grow in an inter-connected mesh of microscopic tubes called mycelium. The cells that make up the mycelium are called hyphae. Somewhat similar to slime-mold plasmodium, hyphae are cells with multiple nuclei; and somewhat similar to filamentous algae, they grow in long strands about 20 times finer than human hair. They grow progressively longer at their tips, and branches form on existing hyphae to form complicated networks. Some mycelium networks can only be described as huge. In fact, a single fungus has been identified that is estimated to cover 900 ha and be about 9000 years old.

            Although fond of edible fungi, I can’t say I’ve ever ordered a Monterey jack and mycelium burger, so where do mushrooms come into this hyphae-delic picture? As I mentioned, mushrooms are a part of the larger fungal organism, specifically the spore-spreading part. Mushrooms are called fruiting bodies and grow above the ground to spread the spores of the underground mycelium. You can think of the fungus as a specialized structure or “organ” of the fungus that has a specific job to do: spread the genetic material of the fungus to a wider area than the mycelium is likely to grow.

Because they grow above ground, mushrooms are also “the face” that the fungus presents to the world. Like any good spokesperson, some mushrooms have an enlightening way of advertising: they glow. The neon green glow, reminiscent of glow-in-the-dark rubber toys, is a result of some very interesting biochemistry called bioluminescence. Bioluminescence literally means light from life, and it is produced by all kinds of life forms, including plants, animals, bacteria, and fungi. The two main players are luciferin (the general name for the group of pigments that are used in bioluminescence), and luciferase (the enzyme helping the reaction along). Like many things that seem pretty magical in biology, bioluminescence is all about chemistry: the luciferase combines luciferin and oxygen to make a high-energy molecule. Like Calvin after one too many bowls of Chocolate Frosted Sugar Bombs, this high-energy molecule is not very stable. It undergoes a second chemical reaction to form a lower-energy molecule, releasing the excess energy as light.

Although not the tastiest of mushrooms (and frequently rather poisonous) bioluminescent fungi have fascinated night-time forest-wanderers for centuries. Their light has been used to mark a luminous path in the woods, and even to illuminate the cabin of the early Turtle submarine. A campfire may make a good s’more, but with Nature’s amazing chemistry there is more than one way to light up the night.

Did Dinosaurs use Celsius or Fahrenheit?

Photo Credit en.wikinoticia.com

As humans there are three ways we’re used to having our temperature taken: under the tongue, in our ears, and the “other” way. Regardless of our personal preferences, we can probably all agree that it is pretty darn difficult to apply any of these techniques to a dinosaur. So barring time machine development, how do we measure the body temperature of our pre-historic pals? Surprisingly, the trick is in the teeth. From molars to metabolism scientists are using some pretty “cool” chemistry to dig up answers.

To start out, let’s talk about metabolism, baby. Our metabolic rate is how fast we break down food, build up body mass, and generate heat. In general, a high metabolic rate means a higher general activity level and higher body temperature. Animals considered to be “warm blooded” have a high metabolic rate and generate their own heat internally, while “cold blooded” animals have a low metabolic rate and get much of their heat from the environment (picture a lizard sunning itself on a rock). This doesn’t mean, however, that “cold blooded” animals always have a lower body temperature, but that their temperatures fluctuate with their environment. In fact, their peak temperatures can be even higher than their “warm blooded” cousins. For this reason, these terms have been replaced by “endotherms” that generate temperature internally, and “ectotherms” that have their body temperature highly dependent on factors outside the body.

Our understanding of dinosaur body temperature has undergone a pretty major makeover over the last two decades. The image of dinosaurs as cold-blooded overgrown lumbering lizards has morphed into a portrait of a much more active and agile group of animals. This increased athletic prowess implies an increased metabolic rate, and sparked the idea that dinosaurs may have been endothermic. Being the investigative creatures that they are, scientists set out to study dinosaur metabolism and body temperature by measuring bone growth patterns, modeling behavior, and studying clues about athletic performance found in footprints. Although many of these studies pointed to an endothermic lifestyle, no real agreement on body temperature was reached.

That was until a group of researchers used the chemistry of dino tooth enamel as a trans-millennial thermometer to accurately measure the average body temperatures of some pre-historic behemoths. Wait just a second, temperature from tooth enamel? How can that work? The answer lies in the power of isotope chemistry. To understand this we have to go down the level of individual atoms of the elements carbon and oxygen. While most atoms of carbon have 12 particles in their nuclei and most oxygen atoms have 16, a very small percentage have extra neutrons and make carbon atoms with 13 particles and oxygen atoms with 18 particles. At high temperatures these “heavy atoms” act pretty much the same as all the other atoms, but at low temperatures they are more likely to bond to each other than one of the lighter atoms. You can think of this like a bad cocktail party: if everyone is really low energy you’re more likely to stick with your friends, but if the party really gets going you're going to meet and mingle with more people. In a technique called “clumped isotope thermometry” scientists measure the proportion of heavy carbon and oxygen that bonded together during tooth formation, effectively measuring the energy level of the tooth-growing party, or dinosaur body temperature.

From analyzing several teeth from large Jurassic Sauropods (“long-necks” to those who remember “The Land Before Time”) researchers calculated their body temperature to be between 36 and 38 ^oC. This is around the same range as most modern mammals. It’s not as simple as whipping out the thermometer from the medicine cabinet, but an inspiring example of how our basic scientific understanding of the world around us can enable scientists to solve seemingly impossible problems. Today dinosaur temperatures, tomorrow, conversations with Neanderthals? Don’t be too quick to rule it out…