Synthetic biology and metabolic engineering are two closely related (and, for the most part, overlapping) disciplines that continue to thrive and expand by the hour. Yet, relatively few efforts tackle the question of expanding the biochemical landscape of cell factories to access truly new-to-Nature products. Pablo Iván Nikel and the SEMsters at the Novo Nordisk Foundation Center for Biosustainability (DTU Biosustain) address this ambitious challenge by constructing synthetic metabolisms in their favorite pet, the soil bacterium Pseudomonas putida
Kostas Vavitsas: Could you tell us about your background and your research journey so far?
Pablo Iván Nikel: I was born and raised in Argentina, and had a soft spot for science for as long as I can remember. Ditto for music, but that is a different story. After graduating as a Ph.D. from the university in Buenos Aires as a biotechnologist and molecular biologist, I decided that it was time for me to explore what you may call «out-of-the-box thinking» as a postdoc. During my Ph.D. thesis, we had been exploring how global transcriptional regulators can be manipulated to boost the production of intracellular biopolymers in Escherichia coli.
Prompted by the systems angle of this applied research, I grew increasingly fond of metabolic engineering ― but most of the examples I could think of were more or less an expansion of traditional molecular biology, that is, «identify the genes encoding enzymes needed to synthesize your favorite product, clone them, throw them in your host, and keep your fingers crossed that your favorite product gets produced!». During a conference in Barcelona in 2009, I had the opportunity to attend a lecture by Víctor de Lorenzo, who would become my advisor during an extended and productive (both from the scientific and personal points of view) postdoctoral stay in his laboratory. Supported by the European Molecular Biology Organization first and then by the Marie Skłodowska-Curie actions of the European Commission, in 2010 I embarked in the journey of understanding and disentangling central carbon metabolism in environmental bacteria. Little I know back then how an exciting ride this would become: we have used a combination of systems and synthetic biology to re-write the core carbon catabolism in Pseudomonas putida (and learnt a big deal along the way!).
Whatever I could say about the importance of engineering the core biochemistry of cell factories is an understatement: perhaps the main conclusion of my work as a postdoctoral associate is that most of the failure of metabolic engineering can be traced to the lack of understanding of how target pathways can be connected to the central biochemistry of the cell. On the other hand, it became apparent that, as complex and clever the approaches could be, metabolic engineering had been focusing on the production of a relatively small number of molecules.
Could authentic novel products be obtained by synthetic biology-guided metabolic engineering? That was my next move as I became a PI. Against Buster Keaton’s advice of going West, I kept moving northward and I accepted a position as group leader of the Systems Environmental Microbiology (SEM) group at the Novo Nordisk Foundation Center for Biosustainability. Since early 2017, I have been assembling a team of awesome collaborators (collectively known as The SEMsters, mind you) and, together, we are expanding the biochemistry of bacteria for the production of new-to-Nature compounds.
Kostas: You are working on engineering synthetic metabolism and producing new-to-nature compounds. What made you choose that research direction and what are the applications of your work?
Pablo: The research in our team has two different (but complementary) angles. On one hand, we are fascinated by the possibility of bringing non-biological chemical elements into life ― that is, to expand the biochemical landscape of living cells. There are some excellent examples of this type of approach in the literature (Nobel laureate Frances Arnold’s impressive work on enzyme evolution comes immediately to mind), in which most of the research has been done in vitro.
We have selected fluorine and other halogens as a model to engineer cell factories for biohalogenation since organic halides have a huge range of practical applications in the industry; yet, they are almost exclusively produced by traditional chemistry. Fluorine is perhaps the best example among the halogens in this regard: one-quarter of all pharma drugs in the market contain fluorine and one-third of commonly used agrochemicals are fluorinated, but all organofluorines are produced by using chemical methods that usually involve highly reactive, difficult-to-manipulate and expensive reagents under extreme physicochemical conditions. In other words, there is an urgent need for sustainable, bio-based approaches for the technical production of organofluorines.
Rather than merely cutting-and-pasting heterologous genes and pathways (which I refer to as «traditional» metabolic engineering) for fluorine incorporation into organic molecules, we are re-writing the core metabolism of environmental bacteria to handle these synthetic biohalogenation reactions. This approach requires the implementation of both bottom-up and top-down approaches and we use multiple approaches for this purpose. Along the line, we aim our efforts at understanding what are the principles that define the boundaries of the observable biochemistry in a cell ― and how can this information be used for engineering synthetic metabolisms for bioproduction.
Kostas: Your organism of choice is Pseudomonas putida. Why did you choose this organism? Do you think the SynBio community should focus on a handful model organisms or expand and use more of the biological diversity?
Pablo: would say that Pseudomonas putida choose me and not the other way around! I got to know the little critter while I was a postdoctoral associate in Víctor’s laboratory and, since then, I have been nothing short of truly impressed by its many peculiarities. For starters, you can find Pseudomonas species in virtually every environment, even (and often!) under very extreme environmental conditions and this is because it has a specialized metabolism.
The metabolic versatility of most Pseudomonas species is amazing: it is a treasure trove to dig for (and engineer) metabolic reactions. This goes hand in hand with a high tolerance to different types of stress: these are the main reasons why we have adopted P. putida as the platform to engineer synthetic biohalogenations, since the reactions and intermediates are either toxic or cause stress to the cells. Very few microorganisms can handle these biochemistries, and P. putida is perhaps the best example (it should be kept very much in mind that this bug has evolved elaborate biodegradation pathways for complex substrates that are usually very toxic). In other words, P. putida is the ideal chassis to engineer for these applications.
However, depending on the purpose, other bacteria (or, more generally, other microorganisms) could be likewise useful. This is still my cri du cœur whenever I am discussing Pseudomonas versus other bacterial species for metabolic engineering and synthetic biology: there is no point to try to adopt a universal chassis simply because there is no such a thing. Yet, if metabolic versatility and stress resistance are desired properties in a bacterial platform, Pseudomonas species are the ideal bugs.
Kostas: Which is your biggest professional or research challenge? (funding, mobility, lack of business partners, something else?)
Pablo: To some extent, the combination of all of them in different amounts is what constitutes a challenge. For instance, doing research in Denmark (and in our center, in particular) is a luxury, and I am not only talking about the financial support that we get from the foundation but also about the stimulating and unique intellectual environment in the center.
Yet, that comes at the price of moving abroad, a challenge that not many people are willing to take (I will write an article about the Danish weather in a different occasion). As a group leader, maintaining the high spirits in your group, creating a sense of community in the team, and keeping the highest research quality standards is a (delightful) challenge that requires a great deal of stamina. Handling rejection is a leitmotiv of academic life, and it is yet another challenge that we all have to deal with sooner or later. In any case, all these hurdles are minor as compared to the positive outcomes: when we discuss science with the people in my group, I immediately forget about the challenges!
Kostas: At the EUSynBioS symposium you lead a breakout session on career development. What is the single most important piece of advice you would give to a Ph.D. student or an early career researcher in your field?
believe in yourself and your ideas, be honest when defining your expectations and have as much fun as possible during the process
Pablo: It was such a pleasure to lead the breakout session on career development during the EUSynBioS symposium, and listening to many different opinions among the attendants. As I did back then during the discussion, I have to disclose that I am a bit biased in my opinions since I simply love working in academia, yet I certainly recognize the value of developing a career in the private sector. In fact, we exploit this angle in our group by collaborating with companies.
If I were to single out a piece of advice to a Ph.D. student or an early career researcher, I would say believe in yourself and your ideas, be honest when defining your expectations and have as much fun as possible during the process. It may sound as an oversimplification, but I am convinced that is actually the essence of making a wise decision for your professional life: what is that you would really like to do as a researcher? How do you react to deadlines and pressure? Are you driven by genuine curiosity? Are you a goal-driven person? These are the essential questions that one should honestly answer when thinking about the professional future.
There is nothing wrong about choosing the academic track or to pursue a career in a company (actually, I get annoyed by the fact that many people tend to see these possibilities as contradicting choices, or mutually exclusive, irreversible ones), as long as this is what you would like to explore (here is where the «having fun» part comes into play). Other than that, imagination is the only actual limit as a scientist (and here is where the «believe in your ideas» part comes into play!).