Playing Evolution: Making Enzymes from Scratch

Photo credit www.jacehallshow.com

In researching almost any topic in biology I am consistently amazed at how natural selection produces things so well suited for their specific roles. From an animal perfectly filling an ecological niche, to an organ efficiently performing a biological task, to a molecule precisely providing the ideal chemistry for its function. It is truly astounding how a process that moves along through random mutation events can result in perfectly coordinated systems at every level. Although natural evolution results in sublimely well-tuned components for every biological system, it also takes a stupendously long time to go about its business. Enter the twenty-first century scientist. While scientists in nanorobotics and materials science are making game-changing leaps and bounds in our ability to manipulate ourselves and our environment, enzyme engineers are starting to give them a run for their money. Have a tough chemical job to do? One that has never been done before? Why not engineer a brand-spanking-new custom enzyme to catalyze your worries away?

            Many of the enzymes that exist naturally are made of protein. The protein itself is a long chain made of different amino acids linked together. You can think of it kind of like a long charm bracelet: the links of the chain form the structure of the chain, while different charms on each of the links make each part of it unique. With proteins, there are 20 types of amino acids or “charms” to choose from that can be strung together in different combinations.

           

            Much like how a ball of wool doesn’t do you a heck of a lot of good on a cold day, a enzyme doesn’t do much at all as one long strand. Mittens or a scarf, however, can be very functional, and so can an enzyme when it is tangled up into a very specific shape. In fact, it is the three dimensional shape that gives a folded enzyme its ability to perform chemistry. So how do enzymes stay folded in the right shape? Going back to the charm bracelet example, some charms (amino acids) can interact specifically with each other to reinforce specific shapes. The “chain” itself also has a tendency to form certain structures: telephone chord-like coils (alpha-helices) and flat sheets formed by the chain doubling back over itself repeatedly (beta-sheets). Additionally, over one third of all proteins contain atoms of iron, zinc, copper, or other metals. By interacting strongly with specific groups of amino acids, these metals can lock part of an enzyme structure in place, helping the rest of the enzyme to also hold its active shape.

            If the interactions that keep an enzyme folded properly and functional seem complex, that’s because they are. A single enzyme has hundreds of individual interactions between individual amino acids and metal groups, some of which can be removed or replaced, and some of which are vitally important either to keep the enzyme in the right shape, or to perform specific chemical functions. Scientists have been trying to tease this apart for years by deleting or swapping out specific amino acids in natural enzymes and observing the effects of these changes. Much like tinkering with an engine vs. building one from scratch, however, you can only learn so much about how something works without creating one of your own. This is exactly what a group of scientists from the University of Michigan reported recently in Nature Chemistry. These intrepid researchers designed an enzyme from scratch: designing a structural “scaffold” (complete with a structure-stabilizing metal) into which they incorporated a small area capable of performing similar chemistry to one of our body’s own enzymes. By testing which enzyme design concepts make the chemistry faster or slower, these researchers are testing and applying the basic principals we currently think govern how all enzymes work.

            Designing enzymes from scratch to perform tasks that natural enzymes already perform is almost the definition of reinventing the wheel. It is, however, incredibly valuable in at least two respects: it demonstrates that we understand how enzymes work well enough to design them from the bottom up, and it opens the door to engineering brand new enzymes that catalyze chemistry not found in nature. Imagine creating an enzyme that quickly breaks down Styrofoam or other long-lasting pollutants. Enzymes are the most powerful chemists in nature, and if we can design their abilities, rather than simply co-opting them, they can be our most powerful industrial and biotechnological tools as well.

Animals on Steroids – Steroids on Oxygen?

In today’s society, the word “steroid” conjures up images of hulking body-builders and sheepish forfeitures of Olympic medals. What most people don’t realize is that steroids actually play a role in all of our lives. In fact, right now there are a whole host of steroids performing the very important role of keeping you alive. Whether we’re pumping up or vegging out, we’re all chock full of steroids. What might be even more amazing is that in addition to keeping animals, plants, and fungi functioning like they should, steroids may also be able to tell us a bit about how and when such multi-celled beings came to be.

            As I alluded to above, steroids aren’t just for greased-up gym junkies. On the contrary, they are enormously prevalent substances produced by all kinds of plants, animals, and fungi. Steroids are molecules made of four rings of carbon fused together in a bent line. This basic structure contains 17 atoms of carbon, and modifying this welded-ring core with additional carbon or oxygen atoms makes hundreds of different steroids. The central steroid in human biology is cholesterol, and it is a vital component of every cell in your body. We also use it to make other steroids like the dreaded hormones that complicate adolescence and control much of our development and body maintenance. While fruit bats, magnolias, and button mushrooms don’t have the same type of awkwardness during puberty, they also produce a preponderance of different steroids nonetheless critical for their own lifestyles.

Cholesterol

            While cholesterol is indeed present in your hamburger, your body also makes it from the simpler molecules you get from digesting all kinds of food. There are three major atomic ingredients in the cholesterol recipe: carbon, hydrogen, and oxygen. The first two come from the food-begotten hydrocarbon building blocks. The necessary oxygen comes from the O₂ in the air we breathe. Like so many things our body makes, the real construction workers are our enzymes. There are many enzymes involved in making steroids, some of which are dedicated to incorporating oxygen from O₂ into the steroid being produced. These enzymes are aptly named “oxygenases”. The job of an oxygenase is not an easy one, and the reaction between O₂ and the hydrocarbons is incredibly slow if no oxygenase is around to speed it along. Many oxygenases use tools like metals and vitamins to manipulate the oxygen into reacting with the carbon in order to come up with the final steroid product.

            Steroids, produced with the help of oxygenases, are one of the major things separating multi-cellular plants, animals, and fungi from life forms made of a single,simple cell, like bacteria. While all “higher” life forms produce steroids, as a rule, bacteria don’t. In fact, steroids are so important to the membranes of more complicated organisms that evolution of everything from venus flytraps to the mailman may not have been possible if our common ancestors couldn’t make steroids. When it comes to steroid production, we need a lot of oxygenases and a lot of oxygen. Way back in the history of life on earth, however, there wasn’t much oxygen floating around in the air like there is today. When there was no oxygen, there were also no steroid-producing oxygenases. As oxygen levels rose, these enzymes evolved, steroids were produced, and in a few hundred million years you were born, went through puberty, and learned all about it. Because of this necessary sequence of events, scientists are able to look at the emergence of steroid production and make connections with changes in the earth’s atmosphere and the evolution of the earliest common relative we share with the coconut.

Studying modern biology can not only help us understand how we are now, but give us insight into where we came from. As for me, the next time someone questions whether an athlete is on the ‘roids, I’ll turn to them and ask, “Isn’t everyone?”

Moonlighting Enzymes: Supporting Keg Parties Everywhere

Our lab is situated in close proximity to a high concentration of fraternity houses, and as the fall term commences the warm autumn atmosphere reeks of fresh loose-leaf, cheap beer, and hang-over bacon. Any truly savvy and overindulgent undergrad should be writing a little thank-you note to one of the crucial factors that enables this year-opening bash: the enzyme alcohol dehydrogenase (ADH). This little microbrew miracle has been helping humans indulge in handcrafted libations for around nine thousand years and is perfectly suited to this function. Or is it? A recent study questions the evolutionary history of ADH and suggests that it might have more important things to do than aid in our enjoyment of an evening martini.

            Given that enzymes are integral components of so much of our body chemistry, it can be hard to remember that they haven’t always been there. Just like everything else, they had to come, to evolve, from somewhere. ADH is no exception. These days, it’s studied as the first enzyme in the metabolic pathway helping us to break down ingested alcohol (ethanol). Without it, we couldn’t efficiently clear alcohol from our systems and it could accumulate to lethal levels over the course of a lively cocktail party. In fact, one form of the enzyme acts in our stomachs, starting to break down the intoxicating molecule before it ever reaches our bloodstream. But where and how did ADH evolve? Were the molecules in our prehistoric ancestors preparing for a time when our species would start breaking out the booze?

            The rules of enzyme evolution parallel those of organism evolution: an ecological (or functional) niche has to exist before the process of natural selection can result in something filling that niche. In the case of ADH this means the availability of ingestible alcohol. If there is no dietary alcohol to break down, there should be no enzyme evolution specifically to break it down. By far, the greatest source of alcohol available for animal consumption is fermentation of fruit sugars, so the appearance of fruit in the prehistoric world should predate that of ADH. This is assuming that ADH really did evolve to break down alcohol. It is this assumption that is being questioned by a group of Mexican scientists.

            In a sobering study examining the functional capacities and evolutionary histories of a range of ADH enzymes, the real purpose of human ADH was called into question. In actuality, humans have several forms of ADH from three very different classes of enzyme using very different means to achieve ethanol transformation. Only one class of ADH is considered to be important for human alcohol breakdown. As it turns out, this class of ADH evolved before the appearance of fruit-bearing plants. That is to say that the type of enzyme we use today to reduce both inhibitions and alcohol toxicity showed up before there was any alcohol to indulge in.

If ADH’s only job was to enable aperitifs, it would not have evolved to do so before the functional niche existed. Instead, it is more likely that ADH has a somewhat serendipitous ability to metabolize the alcohol we drink, and we exploit that ability. In fact, all of the human ADHs are better at catalyzing transformation of different (non-liquor-related) compounds in our bodies, such as bile acids and serotonin. It seems that the party-pleasing activity human ADH is best known for is a handy trick this enzyme would file under “other abilities.”