Which metabolic enzymes should a cell change in concentration to give rise to a large change in steady-state metabolic flux? Which enzymes should an experimentalist inhibit to reduce the flux the most?
Why are some kinase-phosphatase couples in cellular signal transduction ultra-sensitive to changes in signals, while others are not? What is the function of negative feedback in metabolic pathways? How can you design metabolic pathways that are insensitive to particular environmental changes and highly sensitive to others?

Answering such questions requires consideration of enzymes active in networks, as their activity and network influence depend on their reactant concentrations, which are co-determined by all enzymes in the network. Answering such questions therefore needs a systems perspective. One that is quantitative also; as all enzymes influence the network’s functions but to different degrees. The concept of a single rate-limiting step is therefore generally overly simplistic.

Now about 50 years ago, two papers were published, one by Burns & Kacser and another by Heinrich & Rapoport, which dealt with a sensitivity analysis of metabolic pathways to change in enzyme activities and concentrations. They also managed to derive theorems relating sensitivity coefficients. Those coefficients come in two forms: elasticity coefficients and control coefficients, and their products are called response coefficients. We invite you to check out those papers, they have not aged. The entire framework is called Metabolic Control Analysis (MCA).

Bas (Teusink) and myself grew up as young scientists in the scientific community of MCA due to our PhD supervisor, Prof Dr Hans Westerhoff, who was one of the pioneers and advocates of MCA. MCA was therefore part of our training and thinking, and we have published papers about this theory.

What always attracted me (Frank) as how Christine Reder‘s formulation of MCA makes the relation between reaction stoichiometry and enzyme kinetics so clear, and in completely general terms. By using concepts from linear algebra and enzyme kinetics, the control of enzymes on steady-state properties of enzyme networks could be written down in completely general terms. This gave me confidence that a general theory about cellular metabolism and growth can be derived, applicable across all domains of life. Aiming for this has been a research theme throughout my career.

One aspect of Reder’s theory is that the null space of the stoichiometric matrix is used to derive the summation theorems of MCA. But the null space is not unique, which always bothered me when I was a PhD student. This was changed when David Fell, Stefan Schuster and Thomas Dandekar generalised the definition of metabolic pathways in a unique manner, using a concept that is related to the null space of the stoichiometric matrix. They showed that any steady-state flux solution of a stoichiometric matrix is convex combination of a set of unique flux vectors, called elementary flux modes (EFMs). This solved the problem of the entire set of summation theorems (although I do not know whether any one ever published this). EFMs are (beautiful) mathematical objects with extremely appealing mathematical properties for biotechnology and evolutionary biology.

We worked for quite some time on EFMs in the context of the optimisation of stoichiometric models and of dynamic models (containing stoichiometry and enzyme kinetics). It turns that EFMs are the solution of optimisations of enzyme networks, given enzyme kinetics, where one aims to maximise a steady-state flux by optimising enzyme concentrations that sum to a fixed total. They are also the elementary solutions of flux balance analysis computations, considering only reaction stoichiometry and not enzyme kinetics.

If evolution maximises the growth rate of cells then what would be the flux control coefficients of all the optimally expressed enzymes? MCA suggests that you cannot answer this question, because you do not know the enzyme kinetics, and therefore you do not know the elasticity coefficients. But it turns out that you can! This was realised by Klipp & Heinrich, and even earlier by Burn & Kacser in a more simplified setting (in Burn’s thesis). The context of Klipp & Heinrich is regrettably also too simplified to consider the entire metabolic network of a cell that is growing at its maximal rate by having expressed all its enzymes optimally.

The solution to this problem we offer in our paper to the special issue of Biosystems celebrating the 50th anniversary of MCA. You can find the paper here. It is quite a read, we know, but it contains all the main ideas from start to end. We hope that it inspires you to become familiar with MCA, enzyme kinetics, and stoichiometric modelling concepts.

Bacteria grow in communities of many co-occurring species in , e.g., in your gut, in soil, or in the ocean. A fundamental process in these communities (more specifically, communities of heterotrophic bacteria, i.e., bacteria that utilize organic carbon sources) is that bacteria take up substrates (basically, food) like sugars and amino acids from the environment and turn them into biomass or convert them into something else they then excrete. For this new paper, what we wanted to know was: which substrates can different bacteria use (we were focused on substrates they can use as a carbon source)? Can we identify patterns of substrate utilization, e.g., are similar compounds consumed by similar bacteria? Can we predict these patterns by looking at which genes different bacteria encode? Our work touches on several important questions in microbiology, from microbial ecology (how do microbial communities work?) to biochemistry (how does the structure of metabolic pathways shape substrate utilization patterns?) to genomics & evolution (how are capabilities of substrate utilization encoded in the genome, and how did evolution shape these genomic patterns?).

By analyzing the growth of 182 different strains of marine bacteria on 135 different potential carbon sources, we found that we can describe the substrate preferences of our bacteria to a first approximation in terms of their preference for sugars (e.g., glucose or polysaccharides like starch) relative to acids (e.g., amino acids or organic acids which are important intermediate during the chemical conversion of substrates into biomass). This preference is encoded in the genomes of bacteria, which tells us about the evolution of these preferences, but also makes the preferences predictable from genomes.

Our work reveals a way to simplify how we think about the metabolic capabilities of bacteria: we can describe a given heterotrophic bacterium by its degree of specialization along the axis of sugar to acid specialists. This is very useful because it allows us to describe communities of bacteria in a simple way (e.g., by their collective degree of specialization). More fundamentally, our work also shows how the evolution of bacterial genomes is structured by biochemical constraints which drives bacteria to specialize along this axis of sugar to acid specialists. Since the metabolic preferences are encoded in genomes, we can estimate the metabolic capabilities of species that we have not (yet) cultured, but for which we have genomic information (e.g., by sequencing entire communities and piecing together the genomes of the constituent species, a process called metagenomics). This allows us to begin to understand the metabolic processes in many microbial communities in the environment in a simplified manner.

Out now: Genome content predicts the carbon catabolic preferences of heterotrophic bacteria

A custom of our lab is to start the academic year with a state-of-the-union day at the Hortus Botanicus of Amsterdam.

During this day, the PIs give an research overview of the last year, an outlook on the coming year, current duties, and their long-term research vision in the presence of the research (support) staff and all the researchers working on funded projects (PhDs and postdocs).

Bas gave an overview of the Holomicrobiome “groeifonds” project proposal that he is involved in, as an organising member. This is a 240 million euro project that aims to link Dutch microbiome research and company demands: to together drive innovations that will improve the quality of our soils, water, crops and human health by improved methods and technologies. The outlook is that this should give an input to the Dutch economy in an environmentally friendly and sustainable manner. The proposal needs to be reshaped in the coming months, resubmitted, and reevaluated by an independent committee on behalf of the Dutch government for approval (or rejection). Exciting times!

Matti chaired a session on the reconsideration of the logo of our lab, which now has effectively turned this into competition between groups of colleagues. Let’s see what they come up with!

Together with Ralf Steuer (Humboldt-University of Berlin, Germany) we recently wrote a review for Bioessays, see https://doi.org/10.1002/bies.202300015. It addresses how we currently view the consequences of finite biosynthetic resources (for protein expression) for cellular tasks such as stress tolerance, growth and adaptation to new conditions.

We focus on Escherichia coli and Saccharomyces cerevisiae. We acknowledge that the advanced understanding we have of their physiology may not be extrapolatable to microorganisms with a qualitatively different evolutionary history. We argue that these two microorganisms optimise the expression levels of needed metabolic enzymes, given that some fraction of biosynthetic resources is allocated to proteins needed to adapt to new conditions. That latter fraction of non-growth associated protein decreases with cellular growth rate (nutrient quality) and therefore makes fast growing E. coli and S. cerevisiae cells less stress tolerant and adaptive to new nutrients. This is an example of a trade off between growth and adaptation due to finite biosynthetic resources.

If you are interested in such ideas, how they emerge from computational models, and what their precise experimental evidence is then the review we wrote might be of interest to you.

In primate species, a change in lifestyle leads to adjustments in their microbiome. What does this mean? 

Over the last years, ARTIS Micropia Professor Remco Kort and his Bioinformatics & Systems Biology student Isabel Houtkamp studied the faeces of the western lowland gorilla. They did this by comparing the composition of the microbiome of the ARTIS gorillas with that of their wild counterparts – and also with that of humans. They were assisted by Walter Pirovano and Mark Bessem of the company BaseClear, experts in the field of microbial DNA analyses. This research revealed interesting correspondences. In both primate species, a change in lifestyle led to adjustments in their microbiome. So what does this mean? 

To find out, check out their new paper here and a behind-the-scenes story Remco wrote for Micropia.

A new network of over 72 organizations from 12 countries was activated during a convening at ARTIS in Amsterdam on 26–27 September 2022, organized by Remco Kort. Representatives are aligned with the transdisciplinary field and social movement of Planetary Health, which analyzes and addresses the impacts of human disruptions to natural systems on human health and all life on Earth. The new European Planetary Health Hub consists of organizations from various sectors, including universities, healthcare, youth, business, and civil society. The Convening, co-organized by the Planetary Health Alliance (PHA), the European Environment and Sustainable Development Advisory Councils Network (EEAC), and Natura Artis Magistra (ARTIS), aimed to develop Planetary Health Working Groups for Education, Policy Engagement, Research, and Movement Building. The Convening resulted in an outline for each of the Working Group’s aims, visions, missions, priorities, and activities, and set the framework for sustaining their activities in the future through the establishment of the European Planetary Health Hub Secretariat in the Netherlands. The Hub members shared lessons learned, built relationships, and developed artwork-inspired perspectives on Planetary Health. In conclusion, the Convening led to the establishment of a strong European foundation to contribute to the transformations needed for sustainable, just, and equitable societies that flourish within the limits of our ecosystems. The conclusions of the meeting are now published in Challenges.

For more information on the meeting, see VU News and read the meeting summary paper.

Microbial communities play pivotal roles in ecosystems across different scales, from global elemental cycles to household food fermentations. These complex assemblies comprise hundreds or thousands of microbial species whose abundances vary over time and space. Unraveling the principles that guide their dynamics at different levels of biological organization, from individual species, their interactions, to complex microbial communities, is a major challenge. To what extent are these different levels of organization governed by separate principles, and how can we connect these levels to develop predictive models for the dynamics and function of microbial communities?

In a new perspective piece, Matti discusses recent advances that point towards principles of microbial communities, rooted in various disciplines from physics, biochemistry, and dynamical systems. By focusing on principles that transcend specific microbiomes, we can pave the way for a comprehensive understanding of microbial community dynamics and the development of predictive models for diverse ecosystems.

“Searching for Principles of Microbial Ecology Across Levels of Biological Organization” is now published in Integrative and Comparative Biology (https://academic.oup.com/icb/advance-article/doi/10.1093/icb/icad060/7191255)

From June 21-23, Frank is teaching a course at the TU Delft on enzyme kinetics and dynamic models of metabolism. The course information, e.g. the syllabus, can be found here Course information From Enzyme Kinetics to Models of Metabolism.

PhD candidate Maarten Droste is one of the recipients of the famous Red Sock Award for best poster presentation at the SIAM 2023 Conference on Applications of Dynamical Systems in Portland, Oregon. It is a tradition that each prize winner receives a pair of red socks as part of the award. His winning poster, entitled “Determinants of optimal metabolic pathway choice by microorganisms”, is co-authored by Bob Planqué and Frank Bruggeman. 

Maarten’s PhD project is all about understanding the key features and determinants of metabolic pathways choices by microorganisms, using optimality principles and mathematical models.

One of his first results is that physical systems that are alive may obey different principles than those that are inanimate. Whereas inanimate systems generally have increased fluxes (J) at higher thermodynamic driving forces (X) and therefore have a higher entropy production rate (approx. J times X), living systems may adjust their metabolism at higher nutrient concentrations in such a way that the driving force reduces but the flux – which is what matters for cell – still increases, because they swap longer for shorter pathways that only partially degrade the nutrient(s). They do this because enzyme concentration inside cells are bounded and a shorter pathway allows them to have higher enzyme concentration per reaction, and therefore a higher flux. Thus, natural selection of living microbes for growth rate that does not necessarily lead to higher entropy production, as is sometimes stated.

Another one of his findings is that optimal pathway choice depends on the concentrations of nutrients and products of metabolism. And, therefore, on the characteristics of their biotic and abiotic environment.

The abiotic effect is that the nutrient and product concentrations at the cell’s surface matter for the metabolic flux, which are determined by their values at “infinity”, diffusion coefficients, and the rates of cellular metabolism. Since the rates of cellular metabolism contribute to setting those concentrations, metabolic pathway optimization should be done with nutrient and product diffusion incorporated into the nonlinear optimization problem. Maarten has worked out this fundamental problem.

An important biotic effect also plays a role: The concentrations at the cell’s surface are also determined by the activity of other microbial species, and their relative distances, indicating that ecology matters too! Here Maarten has stumbled on a curious finding when the concentration of a product is low enough, a long pathway can have outperform a shorter pathway – even though the long pathway has lower enzyme concentrations per reaction. Whether this happens depends on the kinetics and abundance of nearby microbes feeding on this product. Thus, microbes together shape their “niche”, affecting their optimal pathway choices!

Maarten will be continuing his scientific adventures, wearing his fresh new pair of red sox!

After a few years break, finally it was again time for a lab outing. In the lovely setting of Berg en Dal, we enjoyed three days (April 19th-21st) of networking and conviviality.

“Mens sana, in corpore sano” they say, so we stretched both our muscles, with plenty of biking, jeu de boules, ping pong and football, and also our brains, with the Pub Quiz and Board Games nights.

Multicultural foods and drinks (Italian and Chinese dinners, Dutch lunch and snacks, Lithuanian schnaps,…) provided the right amount of energy.

But there no real fun without some sciency science. We discussed and practised together how to effectively pitch our research interests and ideas. To practise, we split into teams and tried to promote each our own superpowered microbe. (Apparently, cuteness is the best superpower, since the furry Buddy Yeast came out as the winner.)

And, dulcis in fundo, on the last afternoon we crossed the German border – and a sea of sheep – riding the fietstrein!