Scientists capture the elusive structure of essential digestive enzyme (JACS)

Stylized graphic of data on the structure of an active form of an important digestive enzyme, phenylalanine hydolase. The cyan cross-section shows the elution profile and magenta cross-section shows scattering profile. At right is the structure of the activated phenylalanine hydroxylase. Image source: Ando et al.
Stylized graphic of data on the structure of an important digestive enzyme, phenylalanine hydroxylase. At right is the structure of the activated enzyme. Image source: Ando et al.

By Tien Nguyen, Department of Chemistry

Using a powerful combination of techniques from biophysics to mathematics, researchers have revealed new insights into the mechanism of a liver enzyme that is critical for human health. The enzyme, phenylalanine hydroxylase, turns the essential amino acid phenylalanine – found in eggs, beef and many other foods and as an additive in diet soda —into tyrosine, a precursor for multiple important neurotransmitters.

“We need phenylalanine hydroxylase to control levels of phenylalanine in the blood because too much is toxic to the brain,” said Steve Meisburger, lead author on the study and a post-doctoral researcher in the Ando lab. Genetic mutations in phenylalanine hydroxylase can lead to disorders such as phenylketonuria, an inherited condition that can cause intellectual and behavioral disabilities unless detected at birth and managed through dietary restrictions.

Published earlier this month in the Journal of the American Chemical Society, the article presented detailed structural data on the enzyme’s active state – the shape it adopts when performing its chemical duties – that has eluded scientists for years.

“It’s a floppy enzyme which means it’s dynamic,” said Nozomi Ando, an assistant professor of chemistry at Princeton and corresponding author on the paper. “That also means it doesn’t like to crystallize,” she said. This is problematic for the classic method used to study enzymatic structure, known as x-ray crystallography, which requires solid crystal samples. Efforts to crystallize phenylalanine hydroxylase have just recently met success, but still only captured the enzyme in its inactive state.

The researchers in the Ando lab were able to bypass the tricky task of growing crystals of the active enzyme by using their expertise in a special technique akin to crystallography, called small angle x-ray scattering (SAXS), which allows scientists to study enzymes in a solution. And because the enzyme is susceptible to aggregation or clumping up in solution, the researchers coupled their scattering method with a purification technique called size exclusion chromatography (SEC), in which different species in a sample flow through a column at different speeds based on their size.

Steve Meisburger (left) and Nozomi Ando (right)
Steve Meisburger (left) and Nozomi Ando (right)

“Pairing SEC with SAXS is an emergent technique. Our contribution is that we saw a clever way to use it,” Ando said. The experiment is highly specialized and relies on powerful x-rays emitted by particles speeding around the circular track at a synchrotron facility. The research team traveled from Princeton to the Cornell High Energy Synchrotron Source in Ithaca, New York, for multiple intensive data-collection sessions. “Any time on the machine that is available, we use it. Not a single photon gets wasted,” Ando said.

As the enzyme solution passes through the purification technique, flowing across the path of the x-ray beam, researchers record snapshots of the x-ray scattering patterns. The resulting dataset is quite complex as the sample also contains phenylalanine, the compound that “turns on” phenylalanine hydroxylase so that researchers can catch the dynamic enzyme in action.

“Current approaches for analyzing this type of dataset are very crude,” Meisburger said. Essentially, these methods assume that each signal – known as an elution peak – represents a single species, when each peak is actually a mixture of species. In this work, the team used an advanced linear algebra method known as evolving factor analysis that allowed them to separate the scattering components. “We can use these linear algebra methods to ‘un-mix’ species that are overlapping,” Meisburger said, “That’s the piece that I think is really exciting.”

By applying their unique approach, the researchers were able to provide evidence for a model of the active structure of phenylalanine hydroxylase that builds upon recent work by their collaborators in Paul Fitzpatrick’s group at UT Health Science Center at San Antonio. In this model, two phenylalanine molecules dock to a pair of sites on the enzyme, bringing a pair of arms together and freeing up the active sites for doing chemistry once more phenylalanine molecules come along.

“I’m very proud that this is our first paper [published since Ando joined the faculty at Princeton]. We wanted it to be very quantitative and heavy on the biochemistry plus heavy on the physical chemistry. I’m really pleased with the way it turned out,” Ando said.

This work was supported by National Health Institutes grants GM100008 and GM098140 and Welch Foundation grant AQ-1245.

Access the paper here:

Meisburger, S. P.; Taylor, A. B.; Khan, C. A.; Zhang, S.; Fitzpatrick, P. F.; Ando, N. “Domain movements upon activation of phenylalanine hydroxylase characterized by crystallography and chromatography-coupled small-angle X-ray scattering.J. Am. Chem. Soc., 2016, 138 (20), pp 6506–6516.DOI: 10.1021/jacs.6b01563. Published online May 4, 2016.



Theorists smooth the way to solving one of quantum mechanics oldest problems: Modeling quantum friction (J. Phys. Chem. Letters)

Researchers at Princeton
From left to right: Herschel Rabitz, Renan Cabrera, Andre Campos and Denys Bondar. Photo credit: C. Todd Reichart

By: Tien Nguyen, Department of Chemistry

Theoretical chemists at Princeton University have pioneered a strategy for modeling quantum friction, or how a particle’s environment drags on it, a vexing problem in quantum mechanics since the birth of the field. The study was published in the Journal of Physical Chemistry Letters.

“It was truly a most challenging research project in terms of technical details and the need to draw upon new ideas,” said Denys Bondar, a research scholar in the Rabitz lab and corresponding author on the work.

Quantum friction may operate at the smallest scale, but its consequences can be observed in everyday life. For example, when fluorescent molecules are excited by light, it’s because of quantum friction that the atoms are returned to rest, releasing photons that we see as fluorescence. Realistically modeling this phenomenon has stumped scientists for almost a century and recently has gained even more attention due to its relevance to quantum computing.

“The reason why this problem couldn’t be solved is that everyone was looking at it through a certain lens,” Bondar said. Previous models attempted to describe quantum friction by considering the quantum system as interacting with a surrounding, larger system. This larger system presents an impossible amount of calculations, so in order to simplify the equations to the pertinent interactions, scientists introduced numerous approximations.

These approximations led to numerous different models that could each only satisfy one or the other of two critical requirements. In particular, they could either produce useful observations about the system, or they could obey the Heisenberg Uncertainty Principle, which states that there is a fundamental limit to the precision with which a particle’s position and momentum can be simultaneous measured. Even famed physicist Werner Heisenberg’s attempt to derive an equation for quantum friction was incompatible with his own uncertainty principle.

The researchers’ approach, called operational dynamic modeling (ODM) and introduced in 2012 by the Rabitz group, led to the first model for quantum friction to satisfy both demands. “To succeed with the problem, we had to literally rethink the physics involved, not merely mathematically but conceptually,” Bondar said.

Bondar and his colleagues focused on the two ultimate requirements for their model – that it should obey the Heisenberg principle and produce real observations – and worked backwards to create the proper model.

“Rather than starting with approximations, Denys and the team built in the proper physics in the beginning,” said Herschel Rabitz, the Charles Phelps Smyth ’16 *17 Professor of Chemistry and co-author on the paper. “The model is built on physical and mathematical truisms that must hold. This distinct approach creates a new rigorous and practical formulation for quantum friction,” he said.

The research team included research scholar Renan Cabrera and Ph.D. student Andre Campos as well as Shaul Mukamel, professor of chemistry at the University of California, Irvine.

Their model opens a way forward to understand not only quantum friction but other dissipative phenomena as well. The researchers are interested in exploring the means to manipulate these forces to their advantage. Other theorists are rapidly taking up the new paradigm of operational dynamic modeling, Rabitz said.

Reflecting on how they arrived at such a novel approach, Bondar recalled the unique circumstances under which he first started working on this problem. After he received the offer to work at Princeton, Bondar spent four months awaiting a US work visa (he is a citizen of the Ukraine) and pondering fundamental physics questions. It was during this time that he first thought of this strategy. “The idea was born out of bureaucracy, but it seems to be holding up,” Bondar said.

Read the full article here:

Bondar, D. I.; Cabrera, R.; Campos, A.; Mukamel, S.; Rabitz, H. A. “Wigner-Lindblad Equations for Quantum Friction.J. Phys. Chem. Lett. 2016, 7, 1632.

This work was supported by the US National Science Foundation CHE 1058644, the US Department of Energy DE-FG02-02ER-15344, and ARO-MURI W911NF-11-1-0268.

PPPL scientists challenge conventional understanding and improve predictions of the bootstrap current at the edge of fusion plasmas (Physics of Plasmas)

Simulation shows trapped electrons at left and passing electrons at right that are carried in the bootstrap current of a tokamak. Credit: Kwan Liu-Ma, University of California, Davis.
Simulation shows trapped electrons at left and passing electrons at right that are carried in the bootstrap current of a tokamak. Credit: Kwan Liu-Ma, University of California, Davis.

By John Greenwald, PPPL Office of Communications

Researchers at the U.S. Department of Energy’s (DOE) Princeton Plasma Physics Laboratory (PPPL) have challenged understanding of a key element in fusion plasmas. At issue has been an accurate prediction of the size of the “bootstrap current” — a self-generating electric current — and an understanding of what carries the current at the edge of plasmas in doughnut-shaped facilities called tokamaks. This bootstrap-generated current combines with the current in the core of the plasma to produce a magnetic field to hold the hot gas together during experiments, and can produce stability at the edge of the plasma.

The recent work, published in the April issue of the journal Physics of Plasmas, focuses on the region at the edge in which the temperature and density drop off sharply. In this steep gradient region — or pedestal — the bootstrap current is large, enhancing the confining magnetic field but also triggering instability in some conditions.

The bootstrap current appears in a plasma when the pressure is raised. It was first discovered at the University of Wisconsin by Stewart Prager, now director of PPPL, and Michael Zarnstorff, now deputy director for research at PPPL. Prager was Zarnstorff’s thesis advisor at the time.

Physics understanding and accurate prediction of the size of the current at the edge of the plasma is essential for predicting its effect on instabilities that can diminish the performance of fusion reactors. Such understanding will be vital for ITER, the international tokamak under construction in France to demonstrate the feasibility of fusion power. This work was supported by the DOE Office of Science (FES).

The new paper, by physicists Robert Hager and C.S. Chang, leader of the Scientific Discovery through Advanced Computing project’s Center for Edge Physics Simulation headquartered at PPPL, discovered that the bootstrap current in the tokamak edge is mostly carried by the “magnetically trapped” electrons that cannot travel as freely as the “passing” electrons in plasma. The trapped particles bounce between two points in the tokamak while the passing particles swirl all the way around it.

The discovery challenges conventional understanding and provides an explanation of how the bootstrap current can be so large at the tokamak edge, where the passing electron population is small. Previously, physicists thought that only the passing electrons carry the bootstrap current. “Correct modeling of the current enables accurate prediction of the instabilities,” said Hager, the lead author of the paper.

The researchers performed the study by running an advanced global code called “XGCa” on the Mira supercomputer at the Argonne Leadership Computing Facility, a DOE Office of Science User Facility located at the Department’s Argonne National Laboratory. Researchers turned to the new global code, which models the entire plasma volume, because simpler local computer codes can become inadequate and inaccurate in the pedestal region.

Numerous XGCa simulations led Hager and Chang to construct a new formula that greatly improves the accuracy of bootstrap current predictions. The new formula was found to fit well with all the XGCa cases studied and could easily be implemented into modeling or analysis codes.

PPPL, on Princeton University’s Forrestal Campus in Plainsboro, N.J., is devoted to creating new knowledge about the physics of plasmas — ultra-hot, charged gases — and to developing practical solutions for the creation of fusion energy. Results of PPPL research have ranged from a portable nuclear materials detector for anti-terrorist use to universally employed computer codes for analyzing and predicting the outcome of fusion experiments. The Laboratory is managed by the University for the U.S. Department of Energy’s Office of Science, which is the largest single supporter of basic research in the physical sciences in the United States, and is working to address some of the most pressing challenges of our time. For more information, please visit

Read the abstract and article.

The paper, “Gyrokinetic neoclassical study of the bootstrap current in the tokamak edge pedestal with fully non-linear Coulomb collisions,” by Robert Hager and C.S. Chang, was published in the April, 2016, Physics of Plasmas, doi: 10.1063/1.4945615.

Support for this work was provided through the Scientific Discovery through Advanced Computing (SciDAC) program funded by the U.S. Department of Energy Office of Advanced Scientific Computing Research and the Office of Fusion Energy Sciences.