Category: Chemistry

A new open-source toolkit automates the process of computing molecular properties in the solution phase, clearing new pathways for artificial-intelligence design and discovery in chemistry and beyond. The Journal of Chemical Physics published the free, open-source toolkit developed by theoretical chemists at Emory University.

Known as AutoSolvate, the toolkit can speed the creation of large, high-quality datasets needed to make advances in everything from renewable energy to human health.

“By using our automated workflow, researchers can quickly generate 10, or even 100 times, more data compared to the traditional approach,” says Fang Liu, Emory assistant professor of chemistry and corresponding author of the paper. “We hope that many researchers will access our toolkit to perform high-throughput simulation and data curation for molecules in solution.”

Such datasets, Liu adds, will provide a foundation for applying state-of-the-art machine-learning techniques to drive innovation in a broad range of scientific endeavors.

First author of the paper is Eugen Hruska, a postdoctoral fellow in the Liu lab. Co-authors include Emory Ph.D. candidate Ariel Gale and Xiao Huang, who worked on the paper as an Emory undergraduate and is now a graduate student of chemistry at Duke University.

Exploring the quantum world

A theoretical chemist, Liu leads a team specializing in computational quantum chemistry, including modeling and deciphering molecular properties and reactions in the solution phase.

The world becomes much more complex as it shrinks down to the scale of atoms and small molecules, where quantum mechanics describes the wave-particle duality of energy and matter.

Theoretical chemists use supercomputers to simulate the structures of molecules and the vast array of interactions that can occur during a reaction so that they can make predictions about how a molecule will behave under certain conditions. Understanding these dynamics is key to identifying promising molecules for various applications and for driving reactions efficiently.

Researchers have already generated datasets for the properties of many molecules in the gas phase. Molecular properties in the solution phase, however, remain relatively unexplored in the context of big data and machine learning, despite the fact that most reactions occur in solution.

The problem is that studying a molecule in solution requires much more time and effort.

A complicated process

“In the gas phase, molecules are far from each other,” Liu explains, “so when we study a molecule of interest, we don’t have to consider its neighbors.”

In the solution phase, however, a molecule is closely immersed with many other molecules, making the system much larger. “Imagine a solvent molecule surrounded by layers and layers of water molecules,” Liu says. “Depending on its size and structure, a molecule may be covered by tens, or even up to hundreds, of water molecules. In systems of such large size, the computation will be slow and may not even be feasible.”

Before running a quantum chemistry program for a molecule in the solution phase it’s necessary to first determine the geometry of the molecule and the location and orientation of the surrounding solvent molecules.

“This process is difficult to do,” Liu says. “It takes so much time and effort, and it’s so complicated, that a researcher can only perform this calculation for a few systems that they care about in one paper,” Liu says.

Technical issues can also arise during each step in the process, she adds, leading to errors in the results.

A streamlined solution

Liu and her colleagues replaced the complicated steps required to perform these calculations with their automated system AutoSolvate.

Previously, a computational chemist might have to type hundreds of lines of code into a supercomputer to run a simulation. The command-line interface for AutoSolvate, however, requires just a few lines of code to conduct hundreds of calculations automatically.

“The time for running the simulations may be long, but that’s a job for the computer,” Liu says. “We’ve freed the researchers from most of the tedious, manual tasks of data input so that they can focus on analyzing their results and other creative work.”

In addition to the command-line interface geared toward more experienced theoretical chemists, AutoSolvate includes an intuitive graphical interface that is suitable for graduate students who are learning to run simulations.

Labs can now efficiently generate many data points for solvated molecules and then use the dataset to build machine-learning models for chemical design and discovery. AutoSolvate also makes it easier to build and share datasets across different research groups.

Setting the stage for machine learning

“During the past 10 years, machine learning has become a popular tool for chemistry but the lack of computational datasets has been a bottleneck,” Liu says. “AutoSolvate will allow the research community to curate a huge number of datasets for molecular properties in the solution phase.”

Determining the redox potential of a solvent molecule, or the likelihood for an oxidation to occur, is just one example of a key research area that AutoSolvate could help advance. Redox-active molecules hold potential for applications in the development of anticancer drugs and chemical batteries for renewable-energy storage.

“Building up redox-potential datasets will then allow us to use machine learning to look at millions of different compounds to rapidly find the ones with redox potential within the desired range,” Liu says.

Instead of a black-box result, such analyses of large datasets can yield interpretable artificial intelligence, or basic rules for molecular models.

“The ultimate goal is to identify rules that can then be applied to solve a broad range of fundamental science problems,” Liu says.

More information: Eugen Hruska et al, AutoSolvate: A toolkit for automating quantum chemistry design and discovery of solvated molecules, The Journal of Chemical Physics (2022). DOI: 10.1063/5.0084833

Journal information: Journal of Chemical Physics 

Provided by Emory University 

A team of researchers writing in the journal Angewandte Chemie has developed a bioorthogonal molecular system for the targeted introduction of nitrite ions into cells. Their system releases nitrite ions in cancer cells using a “click-to-release” strategy and these ions, along with other active ingredients, help to initiate cell death. The system could improve the synergistic effects of various cancer therapy drugs.

Cells rapidly convert nitrite ions into nitrogen monoxide (NO), which is involved in many cell processes. For example, it can enhance the effect of various cancer drugs by forming reactive oxygen species. However, the targeted introduction of nitrite to a specific location is complicated.

The research groups of Fude Feng at Nanjing University and Shu Wang at the Chinese Academy of Sciences, Beijing, China, have now developed a bioorthogonal system that selectively transports nitrite ions along with other active ingredients to the endoplasmic reticulum, where they are then released.

Bioorthogonal systems facilitate useful chemical reactions (“click reactions”) in cells, without the risk of the reaction partners having adverse effects on the body on their journey to the target site. They have paved the way for an exciting array of novel disease treatment approaches. A testament to this is the fact that the 2022 Nobel prize in chemistry was awarded for the development of click chemistry and bioorthogonal chemistry.

To transport reaction partners to a target site without them participating in unwanted reactions, nitrite ions have to be bound to a carrier molecule as a nitro group. However, the conditions needed to release nitrite again when they reach their target are usually much harsher than those found in living cells. For this reason, the researchers designed two bioorthogonal precursors: one to transport the nitro group and other active ingredients, and another to carry out the click-to-release reaction by reacting with the first precursor.

The first of the two precursors, ER-Non, performed a number of roles. Firstly, it is readily taken in by the endoplasmic reticulum. Not only do many important cell processes take place in this cell organelle, but it is also the site of action of a number of drugs. Secondly, alongside the nitro group, ER-Non transported the active substance novidamide, which triggers cellular stress responses at high doses and can thus cause cancer cells to initiate cell death.

The other molecular precursor, a dithiol, is activated by enzymes typical for cancer cells. In a click-to-release reaction, the activated molecule releases both the nitrite and the novidamide from ER-Non. The chemicals are not simply released; the reaction causes the new substance to fluoresce and, in so doing, to become a photosensitizer.

Under the action of light, it enhances the ability of the nitrite ion and the novidamide to generate reactive oxygen species and to thereby trigger cellular stress. This phenomenon of photosensitizing is used in photodynamic cancer therapy.

The researchers tested their bioorthogonal system on liver cancer cells and observed arrested growth of these cells. They also observed a notable increase in reactive oxygen species after adding both bioorthogonal components. Since none of the components alone would exert this effect, the team concluded that synergistic effects occur. This opens up new possibilities for more effective cancer therapies.

More information: Jian Sun et al, Dithiol‐Activated Bioorthogonal Chemistry for Endoplasmic Reticulum‐Targeted Synergistic Chemophototherapy, Angewandte Chemie International Edition (2022). DOI: 10.1002/anie.202213765

Journal information: Angewandte Chemie International Edition  , Angewandte Chemie 

Provided by Wiley 

Nucleobase-containing coenzymes are believed to be the relics of an ancient RNA world and can provide information on the origin and evolution of proteins. However, coenzyme-protein interactions largely remain unclear.

Recently, researchers from the Okinawa Institute of Science and Technology Graduate University looked at Rossmann enzymes for answers, discovering that insertions and deletions essentially mold coenzyme specificity in these proteins. Their findings, while evolutionarily significant, also potentially provide a novel strategy to engineer coenzyme specificity.

Coenzymes are molecules that assist proteins in nearly half of all the reactions they catalyze. These small, organic molecules contain nucleotides just like in the building blocks of our DNA and RNA. While coenzymes play an extremely crucial role in the catalysis of proteins, their importance is not limited to this alone.

Nucleobase-containing coenzymes are considered the fossil remnants of an ancient RNA-based world, which has been hypothesized to exist even before the very first proteins came into existence. They can, hypothetically, offer a closer look at how proteins emerged and evolved. Unfortunately, not much is known about the evolution of coenzyme-protein interactions.

The structure and function of any protein is coded in its amino acid sequence. Certain structures are evolutionarily conserved across all kingdoms of life. One such recurring structure—the “Rossmann fold”—was discovered by Dr. Michael Rossmann in 1970.

The Rossmann fold is the most catalytically diverse and abundant protein architecture in nature and is an excellent target to study coenzyme-protein interactions. Rossmann proteins display diverse catalytic functions owing to minuscule differences in their structure. These differences, affect the binding specificities of their co-acting catalysts, coenzymes.

Researchers from Okinawa Institute of Science and Technology (OIST) recently modified the coenzyme pocket of a Rossmann protein that naturally performs redox reactions to bind a methylating agent instead. While the natural protein binds to the coenzyme NAD (nicotinamide adenine dinucleotide), the mutant lost the ability to bind NAD and acquired binding for SAM (S-adenosyl methionine). Their findings have been published in Proceedings of the National Academy of Sciences.

Dr. Paola Laurino, assistant professor who leads OIST Protein Engineering and Evolution Unit, says, “As proof of principle, we engineered an oxidoreductase protein to accept a methylating coenzyme SAM instead of its natural coenzyme NAD. This involves the redesign of the ancient and highly conserved glycine rich loop. This task is not trivial because of the complexity of the intramolecular H-bonding and it has attracted the attention of many protein engineers in the past.”

Historically, protein interactions have been studied using site-directed mutagenesis—a process that gives rise to modified proteins where one or more amino acids are substituted with others. However, until now, researchers have not fully unleashed the potential of “insertions” and “deletions” (collectively referred to as “InDels”). In contrast to amino acid substitutions, an “InDel” significantly modifies protein structure because of the addition or removal of one or more amino acids from the corresponding protein sequence.

First, the researchers performed extensive analyses and reshaped the ancient coenzyme-binding motif of NAD into the SAM-binding one. To achieve this, they removed an InDel of three amino acids from the NAD coenzyme pocket and solved the structure of the resulting mutant. As expected, the mutant exhibited the characteristic structural features of a SAM-binding pocket.

Next, the team decided to validate their finding by studying the interactions of the generated mutant with SAM. To this end, the team performed isothermal titration calorimetry measurements—a biophysical technique that determines binding affinities—and validated the successful coenzyme switch when they observed that the mutant was indeed binding. The results were further corroborated through computerized simulations.

Lead author Dr. Saacnicteh Toledo-Patiño, a postdoctoral researcher at OIST Protein Engineering and Evolution Unit concludes, “It is amazing that the sequence combinations possible for a small protein, about 100 amino acids in length, exceeds the number of atoms in the known universe. For this reason, nature has only explored an infinitesimal fragment of these possibilities, and yet, is able to drive the vast number of reactions that sustain life.”

More information: Saacnicteh Toledo-Patiño et al, Insertions and deletions mediated functional divergence of Rossmann fold enzymes, Proceedings of the National Academy of Sciences (2022). DOI: 10.1073/pnas.2207965119

Journal information: Proceedings of the National Academy of Sciences 

Provided by Okinawa Institute of Science and Technology 

The Plant Physiology and Biochemistry research team at the University of Konstanz has discovered previously unknown molecular mechanisms by which plants adapt to their environment—important basic knowledge in times of climate change

Plants are exposed to constant environmental changes, and their survival depends on their ability to sense and adapt to environmental stimuli. Protein molecules in the cell membrane play a crucial role in coordinating extracellular signals and intracellular reactions. The Plant Physiology and Biochemistry research team (University of Konstanz) now succeeded in identifying two deubiquitinating enzymes involved in the molecular mechanism of this adaptation process. The study has been published in the current issue of Nature Communications.

The amount of protein molecules is crucial

In its adaptation process, the cell needs to sense, for example, nutrients or pathogens in its environment. That is the job of special protein molecules—the transporters and receptors, located on the cell membrane separating the inside of the cell from the outside world. They are produced as well as degraded in the cell. Decisive for the plant’s signal perception is the amount of these molecules.

The small signal protein ubiquitin hooks on other proteins and thus ensures that they are degraded. At the same time, there are deubiquitylating enzymes that can reverse this effect by removing ubiquitins.

Biology professor Erika Isono’s team analyzed 18 of these deubiquitylating enzymes in the model plant Arabidopsis thaliana. In cooperation with Karin Hauser, Michael Kovermann and Christine Peter from the Department of Chemistry, the team found two of these enzymes, designated OTU11 and OTU12, which are localized at the cell membrane and are actually involved in regulating the amount of cell membrane proteins.

Lead author Karin Vogel explains the biochemical mechanism: “OTU11 and OTU12 can shorten certain ubiquitin chains. This influences the degradation of the modified proteins.” The biologist also discovered how they are activated: by binding to negatively charged lipids on the cell membrane.

Previously unknown form of activity adjustment

This means that the activity of these enzymes is tightly controlled. Several activation mechanisms for deubiquitylating enzymes are already known. The mechanism reported here is a previously unknown form of activity adaptation. This regulation is so crucial because the effect of deubiquitylating enzymes can have major consequences for the function of the cell.

“Our discovery shows that the deubiquitylating enzymes do not become active until they arrive at the membrane, where the lipids are located. This fits perfectly with the intracellular localization and function of the two enzymes,” says Erika Isono.

Model plants are used in basic biological research to investigate fundamental biochemical and molecular biological mechanisms. The long-term goal is to optimize agricultural yields, which is particularly important in times of climate change, as plant growing conditions can change as a consequence. Erika Isono says, “It’s important that we understand at the molecular level how plants respond to the environment. The ubiquitin-dependent signaling pathway probably plays an important role in this process.”

More information: Karin Vogel et al, Lipid-mediated activation of plasma membrane-localized deubiquitylating enzymes modulate endosomal trafficking, Nature Communications (2022). DOI: 10.1038/s41467-022-34637-3

Journal information: Nature Communications 

Provided by University of Konstanz 

Researchers have investigated the capability of known quantum computing algorithms for fault-tolerant quantum computing to simulate the laser-driven electron dynamics of excitation and ionization processes in small molecules. Their research is published in the Journal of Chemical Theory and Computation.

“These quantum computer algorithms were originally developed in a completely different context. We used them here for the first time to calculate electron densities of molecules, in particular their dynamic evolution after excitation by a light pulse,” says Annika Bande, who heads a group on theoretical chemistry at Helmholtz Association of German Research Centers (HZB). Bande and Fabian Langkabel, who is doing his doctorate with her, show in the study how well this works.

“We developed an algorithm for a fictitious, completely error-free quantum computer and ran it on a classical server simulating a quantum computer of ten qubits,” says Langkabel. The scientists limited their study to smaller molecules in order to be able to perform the calculations without a real quantum computer and to compare them with conventional calculations.

The quantum algorithms produced the expected results. In contrast to conventional calculations; however, the quantum algorithms are also suitable for calculating significantly larger molecules with future quantum computers.

“This has to do with the calculation times. They increase with the number of atoms that make up the molecule,” says Langkabel. While the computing time multiplies with each additional atom for conventional methods, this is not the case for quantum algorithms, which makes them much faster.

Photocatalysis, light reception and more

The study thus shows a new way to calculate electron densities and their “response” to excitations with light in advance, with very high spatial and temporal resolution. This makes it possible, for example, to simulate and understand ultrafast decay processes, which are also crucial in quantum computers made of so-called quantum dots.

Additionally, predictions about the physical or chemical behavior of molecules are possible, for example during the absorption of light and the subsequent transfer of electrical charges.

This could facilitate the development of photocatalysts for the production of green hydrogen with sunlight or help to understand processes in the light-sensitive receptor molecules in the eye.

More information: Fabian Langkabel et al, Quantum-Compute Algorithm for Exact Laser-Driven Electron Dynamics in Molecules, Journal of Chemical Theory and Computation (2022). DOI: 10.1021/acs.jctc.2c00878

Provided by Helmholtz Association of German Research Centres 

What happens in an explosion? Where do the products of that explosion go following the blast? These questions are often difficult to solve. New rugged tracer particles, developed by Pacific Northwest National Laboratory (PNNL) researchers, can provide some answers.

Beyond explosives, many industries may be interested in tracking particulates through harsh environments—which often include high pressures, high temperatures, and different chemicals.

“Lots of chemical tracers exist,” said Lance Hubbard, materials scientist supporting PNNL’s national security research. “The challenge is developing one that can survive harsh environments. It took a few years to convince anyone we could do it.”

Hubbard and his team, along with fellow PNNL researchers April Carman and Michael Foxe, created a tracer that could not only survive but thrive in extreme conditions. Their work was published in MRS Communications.

Quantum dots and water-soaked glass

Organic materials, such as fluorescent dyes, are commonly used as tracers for water leaks and tracking cells in biological experiments. While they work great in those conditions, they aren’t so good for tracing material in explosions. Their problem?

“They burn,” said Hubbard.

Instead, Hubbard and his team focused on inorganic materials to develop their rugged tracers—particularly quantum dots. Though they fared much better than organic materials in harsh conditions, the research team still needed to protect the quantum dots from the extreme conditions of a chemical explosion.

“Finding a way to protect the tracer while still maintaining its luminescent intensity proved to be difficult,” said Carman.

The tracer’s brightness—or luminescent intensity—can be greatly affected by the local environment. Some protective methods can diminish the brightness, making the tracer more difficult to detect. The team focused on using hydrated silica—”basically water-soaked glass” as Hubbard puts it—to protect the quantum dots and maintain their brightness.

Though previous silica coating methods significantly decreased tracer luminescence, the coated tracers designed by the PNNL team were almost as bright as the original quantum dots. Further testing showed that the particles could survive for long periods of time through a range of pH conditions.

“We knew we created something special when we saw our results,” said Hubbard.

Making tracers tunable and mass-producible

Special is one thing, but useable on the commercial scale is another. Lucky for the PNNL team, their synthesis method was designed from the get-go to be completely scalable to produce mass quantities—from kilograms to potential tons per day.

Not only can they make large amounts of the tracer, but they can customize them as well. “We can tune both the tracer’s size and color to any specificity,” said Foxe. “The tracer can be fine-tuned to create a mimic of the mass or material that is being tracked. We can also use a variety of sizes with different colors to visualize how an explosion affects particles of different sizes.”

The tracers are rugged enough to be deployed in harsh environments to track mass and improve scientists’ understanding of environmental fate and transport. They can function under conditions that are too severe for traditional tracers—like in oil and gas refineries or geothermal plants. With tunable parameters and an easy-to-use system, these tracers have many potential applications for tracking material fate and transport in harsh environments.

Persistence pays off

The research has now grown from a small initial investment from the National Nuclear Security Administration (NNSA), Defense Nuclear Nonproliferation Research and Development program to encompassing several related projects.

“We are glad we could keep pursuing this project despite initial skepticism,” said Carman. “We are also thrilled to see where it leads us next.”

More information: Lance Hubbard et al, Luminescent silica microagglomerates, synthesis, and environmental testing, MRS Communications (2022). DOI: 10.1557/s43579-022-00150-3

Provided by Pacific Northwest National Laboratory 

Polychlorinated biphenyls (PCBs) have been widely used in industrial and commercial products including plastics, paints, electronic equipment and insulating fluids. Their manufacture was extensively banned from the late 1970s onwards due to their toxicity, but large amounts still remain in our environment and accumulate inside animals’ bodies.

Chiral PCBs are PCBs that have two mirror-image isomers; these isomers are identical reflections of each other with the same composition. Chiral PCBs are particularly dangerous because they have more chlorine atoms, which are hard for the body to break down, so they can accumulate in the body easily and their isomers are metabolized differently, causing isomer-specific toxicity (particularly neurodevelopmental issues). However, the process behind this selective metabolism was not previously known.

To address this, a research group has illuminated how enzymes produced by the body unevenly metabolize the mirror-image isomers. These results will make it possible to estimate PCB metabolism and detoxification pathways in animals. They will also contribute towards the development of technology to make predictions about chiral PCBs’ mirror isomers, so that we can obtain a better understanding of potential toxicity in humans and other mammals.

The research results were published in Environmental Science & Technology and Chemosphere.

Even though the manufacture and use of PCBs was banned around 50 years ago, they still remain in the environment. It has been discovered that PCBs accumulate inside the bodies of humans and other animals through food consumption. In particular, PCBs with many chlorine bonds are water-resistant and do not break down easily.

This enables high concentrations of these PCBs to accumulate inside animals’ bodies, which adversely affects their health. PCBs’ toxicity is induced by the aryl hydrocarbon receptor (AhR), causing similar adverse effects to dioxin poisoning such as cancer, teratogenesis and immune system damage. Research is being conducted on the particular types of PCB widely known to cause these effects, which are dioxin-like PCBs with one ortho chlorine substitution in the biphenyl ring of their chemical structure, or PCBs with no substitutions.

However, if a PCB has more than three chlorine substitutions at the ortho position of the biphenyl ring, it becomes a mirror-image isomer called chiral PCB. These chiral PCBs do not demonstrate dioxin-like toxicity, but are far more dangerous, binding with the ryanodine receptors (RyR) in organisms to become neurotoxic.

The two mirror-image isomers (called atropisomers) in chiral PCBs have identical physical and chemical properties and exist at a 1:1 ratio in commercial chiral PCBs, but biased ratios are often observed in the environment and in animals such as earthworms and whales, as well as humans. It is believed that this kind of unbalanced ratio is mainly caused by metabolism and that one of a chiral PCB’s atropisomers is more affected by the metabolic reaction, thus reducing its concentration.

Until now, very little research has been carried out into differences in how these atropisomers are metabolized nor the structural arrangement of the metabolic enzymes.

To address this knowledge gap, the team conducted research focusing on the metabolic enzyme cytochrome P450 (CYP enzyme). The CYP enzyme reacts with foreign compounds that enter an animal’s body (for example, chemicals or pollutants in food or medicines). CYP can convert them into water-soluble compounds and promote their expulsion from the body. Previous research by this group has shown that CYP enzymes hydroxylate and dechlorinate dioxin-like PCBs.

This decreases a PCB’s binding with AhR and increases its water solubility, promoting expulsion from the body and therefore counteracting its toxicity. In other words, CYP is an important enzyme that determines whether or not PCBs are treated as toxic compounds by the body. To measure the metabolic action of CYP on chiral PCB, the researchers set up a CYP enzyme and PCB docking model. They used this to estimate the structure of PCB metabolites and the structure of the CYP that decides to metabolize each of the PCB atropisomers differently.

For the experiment, the group selected three types of chiral PCB, each with a different number of substituted chlorine atoms; CB45 (4 chlorine substitutions), CB91 (5 chlorine substitutions) and CB183 (7 chlorine substitutions). They separated the atropisomers for each type of chiral PCB using chromatography and let them react with a human CYP enzyme. It is believed that research on separating the atropisomers and letting them react has not been done before now.

The results revealed big differences in how each atropisomer is metabolized. Even though the two atropisomers in one PCB have the same physical and chemical composition, they are biologically different. The researchers found that one of the chiral PCB atropisomers was metabolized more than the other one, disrupting the 1:1 ratio.

In addition, it is thought that the amount of (aS)-CB183 atropisomer decreases because it is metabolized more than the other atropisomer, and this is supported by the reports of low accumulation of (aS)-CB183 in humans.

But why are these physically and chemically identical atropisomers metabolized differently by the CYP enzyme? To solve this mystery, the researchers used a computer model to investigate how easily each chiral PCB atropisomer binds to the chemical structure of CYP. They found that when an atropisomer fills up the substrate-binding cavity inside the CYP enzyme, CYP’s amino acids (that form the cavity) interfere with the binding between CYP and the atropisomer.

Therefore, the atropisomer that isn’t interfered with by CYP’s amino acids becomes easy to metabolize (atropisomer (aR)-CB45 in CB45, and (aS)-CB183 in CB183), resulting in alterations to the original 1:1 ratio of atropisomers found in chiral PCB.

The results of this research will be useful for making predictions about the atropisomers of chiral PCBs, which accumulate easily inside animals’ bodies. In other words, it will be possible to work out which atropisomer is reduced by the metabolic reaction with CYP enzymes and which atropisomer remains inside the body. A chiral PCB’s toxicity is activated by binding with RyR, though the ability to bind with RyR differs between the atropisomers. Therefore, this research will make it possible to estimate the toxicity of chiral PCBs.

More information: Hideyuki Inui et al, Differences in Enantioselective Hydroxylation of 2,2′,3,6-Tetrachlorobiphenyl (CB45) and 2,2′,3,4′,6-Pentachlorobiphenyl (CB91) by Human and Rat CYP2B Subfamilies, Environmental Science & Technology (2022). DOI: 10.1021/acs.est.2c01155

Terushi Ito et al, Enantioselective metabolism of chiral polychlorinated biphenyl 2,2′,3,4,4′,5′,6-Heptachlorobiphenyl (CB183) by human and rat CYP2B subfamilies, Chemosphere (2022). DOI: 10.1016/j.chemosphere.2022.136349

Journal information: Chemosphere  , Environmental Science & Technology 

Provided by Kobe University 

Tropane alkaloids are a particular class of plant-derived compounds that have been exploited by mankind since the domestication of medicinal plants. The distribution of these alkaloids is scattered amongst the flowering plants and the two most studied families include those from the Solanaceae (tomato, tobacco, potato relatives) and the Erythroxylaceae (coca). The WHO lists several tropane alkaloids as some of the most important medicines in the modern day pharmacopeia. However other compounds such as cocaine are more infamous for their narcotic and euphorigenic properties.

“It is critical to understand how plants produce these alkaloids in order for mankind to continue to build upon nature and develop new useful medicines,” says Dr. John D’ Auria, head of the IPK’s research group “Metabolic Diversity.”

The most studied and characterized system for tropane production has historically been within Solanaceae. There are more than ten chemical modification steps necessary to transform the beginning amino acid precursors into the final active alkaloids, and all of these steps have been identified and characterized in solanaceous plants.

The scattered distribution of tropanes among flowering plants has always hinted that different families may have developed the ability to produce these alkaloids independently from one another. In fact, several steps of tropane biosynthesis were already documented to have evolved independently within members of the Erythroxylaceae.

“We have been working on elucidating the coca-derived tropane pathway for the last 15 years and we have been successful in working on several key steps in the biosynthesis of cocaine and other related tropanes in coca,” say the researchers. “The idea that coca would share similar enzymes and genes with their distant solanaceous relatives was incorrect. While the final structure of tropanes is similar, the pathway leading to these alkaloids is different.”

In order to discover the last remaining steps of the pathway in coca, Dr. D’ Auria collaborated with the lab of Dr. Christina Smolke from Stanford University. The Smolke group are experts at manipulating yeast and microorganisms to produce important medicinal compounds via synthetic biology methods. Their combined research is published in the journal Proceedings of the National Academy of Sciences.

“With [the Smolke group’s] assistance, we used the multiplicative power of gene manipulation in yeast to test many different gene candidates for the missing steps in the coca pathway. In essence, at every unknown step, we designed and tested multiple candidate sequences,” the researchers report.

These candidate sequences originated from transcriptome studies performed in Dr. John D’ Auria’s group as well as the group of Dr. Lyndel Meinhardt from the USDA in Beltsville, Maryland (U.S.).

“Using this powerful gene discovery platform, we successfully identified all the remaining ‘missing steps’ for tropane biosynthesis in coca. This represents the culmination of more than ten graduate student projects in my group and 15 years of my research,” says Dr. D’ Auria.

The most significant portion of the findings now confirms that tropane biosynthesis has independently evolved at least twice during the evolution of flowering plants. “This is important because we also show in our study that you can mix and match the Solanaceae and Erythroxylaceae genes and produce tropanes,” say the researchers. In layman’s terms, the research provides multiple tools for synthetic biologists to begin designing the tropane alkaloid pathway in organisms that have never produced them before, and with the ability to use different enzymes for similar steps, it is possible to optimize or modify those steps for specific chemical outcomes.

“In addition, we also show that the beginning portion of the pathway in coca proceeds by an interesting ‘detour’ or alternate route that doesn’t exist in solanaceous species,” says Benjamin Chavez, the first author of the study and a Ph.D. student in the D’Auria laboratory. “This provides insights in how plant metabolism can find solutions to biochemical challenges. Namely, we can understand the interplay between early precursors and their bottlenecks.”

Lastly, the researchers discovered a specific enzyme that is responsible for the so-called “carbomethoxy group” present exclusively in coca alkaloids. Solanaceous species do not have this modification. The carbomethoxy group is partially responsible for the euphorigenic properties of cocaine.

More information: Elucidation of tropane alkaloid biosynthesis in Erythroxylum coca using a microbial pathway discovery platform, Proceedings of the National Academy of Sciences (2022). DOI: 10.1073/pnas.2215372119

Journal information: Proceedings of the National Academy of Sciences 

Provided by Leibniz Institute of Plant Genetics and Crop Plant Research

You can keep your best guesses. Engineers at Rice University’s George R. Brown School of Engineering are starting to understand exactly what goes on when doctors pump contrast agents into your body for an MRI scan.

In a new study that could lead to better scans, a Rice-led team digs deeper via molecular simulations that, unlike earlier models, make absolutely no assumptions about the basic mechanisms at play when gadolinium agents are used to highlight soft tissues.

The study led by Rice chemical and biomolecular engineer Philip Singer, former associate research professor Dilip Asthagiri, now of Oak Ridge National Laboratory, and graduate student Thiago Pinheiro dos Santos appears in Physical Chemistry Chemical Physics.

It employs the sophisticated models first developed at Rice for oil and gas studies to conclusively analyze how hydrogen nuclei at body temperatures “relax” under nuclear magnetic resonance (NMR), the technology used by magnetic resonance imaging, aka MRI.

Doctors use MRI to “see” the state of soft tissues, including the brain, in a patient by inducing magnetic moments in the hydrogen nuclei of water molecules to align with the magnetic field, a process that can be manipulated when gadolinium agents are in the vicinity. The device detects bright spots when the aligned nuclei relax back to thermal equilibrium following an excitation. The faster they relax, the brighter the contrast.

Gadolinium molecules are naturally paramagnetic and sensitive to magnetic excitation. Because they’re toxic, they are usually chelated when part of a contrast agent. “A chelate basically hugs the gadolinium and protects your body from directly interacting with the metal,” Pinheiro dos Santos said. “We’re asking, exactly how do these molecules behave?”

Though gadolinium-based contrast agents are injected by the ton into patients each year, how they work on a molecular level has never been fully understood.

“Going back 40 years, in the NMR field people assumed liquid water is just a collection of marbles moving about, and the dipoles in the marbles randomly reorient,” Asthagiri said.

But such assumptions are limiting, he said. “What Thiago does with his explicit simulation is show how the water network evolves in time,” Asthagiri said. “These are complicated, computationally intensive calculations.”

The Rice simulations make use of highly refined, polarizable force fields to study the phenomenon in detail, and that required intensive GPU-accelerated computing.

The team validated its molecular dynamics approach with experimental data by co-author Steven Greenbaum, a professor of physics at Hunter College in the City University of New York, whose lab specializes in NMR measurements of ionic and molecular transport processes in condensed matter.

The simulations revealed distinct differences in how the inner and outer shells of water molecules around gadolinium respond to thermal excitation. “The inner shell is the group of eight or nine water molecules around gadolinium,” Pinheiro dos Santos said. “They’re strongly attached to the gadolinium and they stay there for a long time, a few nanoseconds. The outer shell encompasses all of the remaining water molecules.”

The researchers found that while the structure of the inner shell does not change between 41 and 98.6 degrees Fahrenheit, its dynamics are very susceptible to thermal effects. They also discovered that temperature greatly affects the self-diffusivity of molecules in the gadolinium-water simulations in a way that affects outer-shell relaxation.

“Overall, these discoveries open a new way to elucidate how contrast agents respond at human body conditions during an MRI scan,” Singer said. “By better understanding this, one can develop new, safer and more sensitive contrast agents, as well as use simulations to enhance the interpretation of MRI data.”

He said future studies will examine chelated gadolinium complexes in fluids that are more representative of cellular interiors.

More information: Thiago J. Pinheiro dos Santos et al, Thermal and concentration effects on 1H NMR relaxation of Gd3+-aqua using MD simulations and measurements, Physical Chemistry Chemical Physics (2022). DOI: 10.1039/D2CP04390D

Journal information: Physical Chemistry Chemical Physics 

Provided by Rice University 

Benzobactins are bacterial natural products that have special biological activity due to a compound consisting of two ring structures. The bacterial genes responsible for the formation of the compound were previously unclear. Now, scientists at the Max Planck Institute for Terrestrial Microbiology have been able to decipher its biosynthesis through extensive genomic research. Their research facilitates the discovery of numerous previously unknown natural compounds for medical drug therapy.

In their natural habitat, microorganisms are often exposed to changing environmental conditions that require numerous survival responses. The most efficient one is their capability to produce a wide array of natural products with diverse chemical structures and functions.

Benzobactines—powerful, but rare

Benzoxazolinate is a rare natural compound that confers extraordinary bioactivities on natural products. It is, for example, the essential part of lidamycin, an antitumor antibiotic that is one of the most cytotoxic compounds so far. The reason for this capacity is the fact that benzoxazolinate consists of two rings, a structure that allows it to interact with protein as well as with DNA. However, tracking down the producers of this rare substance in nature resembles the proverbial search for a needle in a haystack.

In order to exploit new pharmaceutically valuable natural compounds, like antibiotics, tumor suppressants or immunosuppressants, it is necessary to know the responsible genes, or more precisely, their biosynthetic gene clusters (BGCs). BGCs are locally clustered groups of two or more genes that together encode the production of a certain set of enzymes—and thus the corresponding natural products produced by these enzymes.

So far, the biosynthetic gene cassette of benzoxazolinate remained elusive, hindering to expand the repertoire of bioactive benzoxazolinate-containing compounds. More specifically, the last formation step of benzoxazolinate was unclear. Now a team of Max-Planck scientists led by Dr. Yi-Ming Shi and Prof. Dr. Helge Bode succeeded in the biosynthetic characterization of the benzoxazolinate pathway.

During the biosynthesis, the pathway obviously “borrows” an intermediate from the so-called phenazine pathway, responsible for the production of another natural product. Most importantly, the researchers identified the enzyme that is responsible for the last step, the cyclization towards benzoxazolinate.

Using an enzyme as a probe for natural substances

Ph.D. student Jan Crames, co-first author of the study, explains, “Knowing the enzyme’s identity, we used it as a probe. With genome-mining we were able to detect many closely related biosynthetic pathways for benzoxazolinate-containing natural products, so-called benzobactins.”

According to the scientists, the most striking aspect was the wide distribution of these genes in other bacteria. “These pathways were found in taxonomically and ecologically remarkably diverse bacteria ranging from land to ocean, as well as plant pathogens and biocontrol microbes. Their wide distribution indicates that these molecules have a significant ecological function on the producers,” as Yi-Ming Shi, first author of the study indicates.

Prof. Helge Bode, leader of the department “Natural Products in Organismic Interactions” at the Max-Planck Institute for Terrestrial Microbiology in Marburg, adds, “Our findings reveal the immense biosynthetic potential of a widespread biosynthetic gene cluster for benzobactin. Now, we have to find out their ecological function and if we can apply them as antibiotics or other drugs.”

The research was published in Angewandte Chemie International Edition.

More information: Yi‐Ming Shi et al, Genome Mining Enabled by Biosynthetic Characterization Uncovers a Class of Benzoxazolinate‐Containing Natural Products in Diverse Bacteria, Angewandte Chemie International Edition (2022). DOI: 10.1002/anie.202206106

Journal information: Angewandte Chemie International Edition 

Provided by Max Planck Society