Hi, my name is Jason Bock, and I'm an Assistant Professor in the Department of Chemical, Paper and Biomedical Engineering in the College of Engineering and Computing at Miami University. In today's video, I'm going to talk to you a little bit about the research that I have ongoing at Miami, in metabolic and protein engineering, as well as give you a sense of some of the problems within these fields in our solutions for them. If you'd like to summarize my research in one slide, it's using nature as an inspiration to come up with engineering solutions. So because nature has had several millennia of evolution, ahead of when we started even started doing any engineering, there are many solutions that nature has already come up with that are more clever than what we can design right now. My job is to go through and find what those solutions are, characterize them, and then apply them to various fields within engineering to benefit society. And so I'm going to give you a sense of how I can go through nature and find these solutions through both metabolic and protein engineering projects. When I think about metabolic engineering and its potential, I start with the sort of renewable bio production. And this is one of the key aspects of metabolic engineering and why I think it has a huge upside over conventional chemical synthesis. And the reason that it's renewable is because we're taking sunlight and carbon dioxide in fixing them into plants, such as sugar beets that I show you on the lower left hand side, we can then take these sugars and convert them using microbes into a variety of different products. And I'm going to talk about those products on the next slide. Because of this renewable nature of this, it's often more desired than say, using fossil fuels, which are commonly used in the chemical synthesis market right now. And so the key aspect of metabolic engineering is the fact that we're going to re re wire the metabolism of bacteria or microbes, I should say, and in my case, bacteria, so that we take it from where it usually takes these sugars and just makes more cells with them, we're going to make a variety of different products. And that's the other key aspect of metabolic engineering is the fact that we can make a wide variety of biomolecules. And that's because cells contain a set of diverse catalysts called enzymes that can convert sort of substrates like those sugars into all of the different molecules that you see here, right in each one of these enzymes will perform a very specialized function. And often that gives us things that conventional chemistry cannot do because these enzymes from nature can do chemistry that we haven't even discovered yet in sort of laboratories using conventional catalysts. The other nice thing about doing all of these inside of cells is you don't use harsh chemicals to do it. Because if you're using harsh chemicals, it's going to kill your your cells. So all of those are some potential advantages to going the metabolic route versus conventional chemical synthesis. So to give you a sense of some of the areas impacted by this, so for instance, one three propane dial is a commodity chemical that's used, made by DuPont and is used in making carpet fiber isobutanol is a short chain biofuel. So to replace conventional gasoline, lycopene is actually the red coloring in tomatoes, but it also has some antioxidant properties. So it's a nutraceutical molecule, artemisinin is a precursor to a malarial drug. So something to be used in the healthcare industry. vanillin is a molecule that gives the vanilla flavor that you've had when you bake cookies. And violation is a purple dye that's also used as an antimicrobial sort of molecule. So you can see that there's a huge set of chemicals that we can make here. And this is just a tiny fraction of what we use metabolic engineers can do. And you can see that, you know, the structure of each one of these chemicals will determine maybe what its purpose is in the chemical industry and how we make it.
So before we start talking about how we make the enzymes, I'm going to talk about sort of, you know, how do we get different enzymes inside of stem cells? So as I said, right, we're going to take cells that naturally have a metabolism, right? We're going to rewire it to make those different molecules that we want to make, right? We're going to do that by adding different enzymes to do it. But so how do we get those enzyme enzymes inside of the cell? And we start with the central dogma of biology for this, in this central dogma of biology basically states that DNA right is transcribed into mRNA, which is then translated into protein, right in protein is another way or is a string of amino acids, right. And those amino acids fold into a structure in one class of those proteins is enzymes, which is what I was just talking about before. Okay, so how can we get the enzymes inside of cells. And the way that we do this is actually by getting DNA inside of cells, because DNA is the universal genetic code. So basically, the DNA from one organism can be read in another organism. And so let me give you an example of this. So if we look here, we have a picture of a jellyfish. And this jellyfish contains a protein called green fluorescent protein. And that protein allows the jellyfish to actually glow green. Okay, and so this is the sort of protein right, so this is the end of the central dogma of biology that allows for the function of the green and the jellyfish. Now, say I want to make bacteria glow green. Well, bacteria normally don't do that. Right? So what do I have to do? Right? I can't take the protein and put it into the bacteria, right? Rather, what I can do is I can find what is the DNA sequence that encodes for this protein, this green fluorescent protein, and I can then insert that DNA sequence into the bacteria, right, then that bacteria can go through the central dogma of biology of transcribing, and translating, resulting in green cells. And so these are a picture of some cells that I had imported the gene for green fluorescent protein into, and they were able to glow green. Okay. So that's kind of fun that we can sort of change sort of the sort of the ability of these cells to do these fun functions. But can we actually use this in an engineering way. And so here's another project that I had worked on, where I took a gene from lactococcus lactis, which you commonly known as the bacteria that you find in yogurt, right? And this gene is able to perform a function where it takes an amino acid intermediate and can convert it into the isobutanol molecule that you see over here. So again, how did I do this, I took the gene from the the lactococcus, lactis, right, that gene encodes for the the enzyme structure that you see here, notice that this enzyme looks very different than the green fluorescent protein, right in that structural differences, what allows to have that different function. And the reason it has the different structure is because it has different DNA, right, and I put that different DNA into the cells. And then the cells are able to make this isobutanol molecule, which as we said before, a potential biofuel replacement. Okay, so so this is kind of the heart of what we're doing right is we're moving genes from one organism to another, to perform these unnatural functions. And you can see that where nature comes in, right, as nature is providing us with all of these different sequences that we can start looking through, to find the ones that provide the function that we're looking for.
And so our approach in my laboratory is twofold. So we look for two different things. We're looking at building different pathways in finding different novel organisms to put these in, right, I'm going to describe two projects within that space, and how we're approaching this pathway building. Because while it's easy to put an enzyme inside of a cell, it's difficult to get it to interact with native metabolism in a way to make the chemicals that you want, in a high amount, right. So the goal of making these chemicals is to make enough of it so that we could supply it sort of into the the sort of world supply to meet the global demand, right. And so to do that, right, we have to come up with clever ways of say, making the enzymes at different times or making them in different amounts. And so that's the pathway building. And we can put them in different organisms that might have some unusual traits that will allow us to do some easier bio processing in the long run. And so one of the projects that we're working on, is looking at these transcription factors called sigma factors that we can use within metabolic engineering. And so the Sigma factor is a class of molecules that allows RNA polymerase. So this is the part of the central bog dogma that's doing the transcription step, right? To interact with a minus 35 and a minus 10 position. So these are base pairs upstream of say, the gene for the enzyme that you're looking to make in your metabolic pathway, right? What we're trying to do is take sigma factors from one organism and placed them in another organism. And what we can do with that similar to the genes that we talked about before, is now we can use these as a control knob to basically you can Roll when our pathway is being made. And so you can see this on the right hand side of this slide, where we use this alternative sigma factor, right to make these different enzymes, which I cleverly show is Pac Man. And these enzymes right can then come together to constitute a metabolic pathway where we say, convert a red triangle into a purple hexagon. And we can do that by controlling that trend when each one of these genes and then enzymes is transcribed. So we think that this is a clever way to potentially increase the amount of bio production that we can get a nice house. Another project that I've been working on is actually engineering environment, environmentally isolated bacteria. And this environmental isolate comes from an underground carbon dioxide reservoir in Colorado. So one of my colleagues went out and found sort of some organisms that grew there. And these organisms can uniquely survive under really high pressures of carbon dioxide. And we're going to use that in some bio, bio prior, we're going to use that as an extractive solvent. So that we can make some chemicals and extract them into the the carbon dioxide. And we think that this is a clever way to use a novel organism and provide some biotechnological benefit. So while putting together enzymes into pathways is a really interesting and useful engineering tool. I'm also interested in something called protein engineering. And so when protein engineering on like metabolic engineering, it's all about the proteins here, right. And these proteins, we've already seen one example them and those are enzymes, right? So enzymes are those biological catalysts that we talked about before, right. But there's other whole classes of enzymes or other proteins out there, such as antibodies, which are recently in the news. Because there have been some that have been developed to help stave off some of the, if you got infected with COVID, they can help mitigate some of the symptoms of that. And then there's other ones that we can use as different biomaterials. So here's a picture of something called a lesson. And this is an extracellular matrix protein that we can use, it's actually stretchable. Right. So we can use that that property for making some say, wearable electronic devices. And so here, what we can do, so one of the things we can do is engineer enzymes, right to make them better catalysts, right, so we can put them directly into into pathways that we just talked about. But there's some other uses for these too. And I think that my favorite one to talk about is in a laundry detergent. Right? So laundry detergents, one of the widest uses of enzymes, industrially, right, and there's a lot of different enzymes in there. And those enzymes, what they're doing is they're actually able to break down your different stains, so maybe a grass stain, or if you've got, say, some some fat on your, on your clothes, right? They'll break those down, right? Or other sort of protein stains, right? So what these enzymes can do is, right, it's going to help you make your laundry detergent better. So how could we maybe make some better catalysts for this? Right? We could make them faster, right? So that would be something that would be very useful so that you can break down more stains, maybe we could make them more reusable, right? So that you wouldn't have to buy laundry detergent, every time maybe we can make them sort of go to where the stain is, can we can we target them to the stains? Right? All of these things are what the goals of protein engineering is, is can we make these catalysts? Fun? Or can we make these catalysts better? And can we then put them into different products to really benefit from the enhancement of the engineer protein? So where I get interested in this field, is in looking through what nature has already made for enzymes? So for instance, if we have an enzyme, right, we know that it's encoded by DNA, and there's a huge sequence of A's, t C's and G's that do this, right? This is only maybe the part of a, an enzyme sequence. Right? And while that looks like gibberish to us, right nature has found ways to them make these into these exquisite catalysts that we talked about before with really high specificity. And I get interested in trying to find other enzymes from nature that are related to the ones that we currently use, right in studying the different properties that they might have in the hope that we can sort of find ones that have slightly altered properties and maybe these are altered in a good way, maybe they are faster than the other one was, maybe it allows us to do a slightly different chemistry than it could before. Right? And we can sort of put together these maps. So say here we have in light blue an enzyme that we know works really well. And then we can look at other variants of this enzyme and how related they are to each other so that we can start going through mining the sequence space, and finding variants that potentially could be useful in biotechnology. Okay, so with that, I hope I've given you a little bit of a sense of, you know, how I approach some problems at Miami University. Some of the outstanding issues within metabolic and protein engineering, as well as some of the fun ways that nature can be used to inspire us as engineers.