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Structure of Human Phenylalanine Hydroxylase Provides Insight Into Possible Treatment Strategies for PKU

August 27, 2019

Although once thought to be anomalous, the basic science driving Jaffe’s work is now realized to be pertinent to proteins whose dysfunction contributes to cancer. It has also provided a “shape-locking” approach to new therapeutics. Although once thought to be anomalous, the basic science driving Jaffe’s work is now realized to be pertinent to proteins whose dysfunction contributes to cancer. It has also provided a “shape-locking” approach to new therapeutics.

PHILADELPHIA (August 27, 2019) – New insights from researchers at Fox Chase Cancer Center into the structure of full-length human phenylalanine hydroxylase (PAH) may aid ongoing efforts to develop effective treatments for the metabolism disorder phenylketonuria (PKU).

PKU is an inherited error of phenylalanine metabolism caused by deficiency in the enzyme PAH. Infants are tested for PKU, which is diagnosed in approximately 1 in 10,000. People with PKU must maintain a diet low in phenylalanine, which is found in most protein-containing foods such as milk, chicken, beef, pork, nuts, and soy. Undiagnosed or untreated, PKU can cause growth failure, seizures, mental disability, and organ damage.

“When phenylalanine concentrations start to rise, like after someone eats a steak for example, they can rise to toxic levels,” said Eileen K. Jaffe, PhD, a professor in the molecular therapeutics program at Fox Chase. “Resting PAH needs to have a way to turn on. It does this through dramatic changes in assembly architecture.”

More than a decade ago, Jaffe identified shape-shifting proteins like PAH, whose normal physiological function requires major changes in protein assembly architecture and for whom defects in these dynamics contribute to disease. Although once thought to be anomalous, the basic science driving Jaffe’s work is now realized to be pertinent to proteins whose dysfunction contributes to cancer. It has also provided a “shape-locking” approach to new therapeutics. 

More recently, Jaffe and her team solved the first X-ray crystal structure of full-length PAH. This structure consists of an asymmetric arrangement of four identical subunits, known as a tetramer, in a low-activity form, or resting form, that predominates at low levels of phenylalanine. Additional solution studies show that a distinct four-unit structure characterizes the activated form of PAH.

The evidence that alternate forms of PAH are represented by distinct tetramer structures supports Jaffe’s recently proposed model for the activation of PAH by dietary phenylalanine. As phenylalanine levels increase, the structural equilibrium of PAH shifts toward its active assembly, allowing for the metabolism of phenylalanine. Once phenylalanine is metabolized, the protein converts to its resting state.

In people without PKU, the protein switches from a resting state to an active assembly when phenylalanine reaches levels of > 50μM. For some people with PKU, this switch occurs at a much higher phenylalanine concentration, resulting in levels well above a neurotoxic threshold.

These findings were outlined in a recent paper, “Biophysical Characterization of Full-length Human Phenylalanine Hydroxylase Provides a Deeper Understanding of its Quaternary Structure Equilibrium,” which was published in The Journal of Biological Chemistry.

In that paper, Jaffe and her team described the human PAH structure in its resting-state conformation and refined the best working model of the activated conformation. The latter may prove particularly valuable in the development of therapeutics designed to shape-lock this conformation.

During their research they designed a variant of human PAH called C29S that allowed them to get a crystal structure of the resting state of the protein. Comparison of this human crystal structure with a previously determined rat structure revealed that parts of the human protein moved much more, particularly a part that dramatically relocates during the activation process.

“Understanding where and how the human protein moves is a useful piece of information because it can help us understand how different disease-associated variants might affect the protein,” Jaffe said.

Jaffe theorizes that the reason there are so many disease-associated variants in PKU is that any changes in the stability of the various protein structures, or conformations, can shift the equilibrium between the resting-state and activated forms, the latter of  which is stabilized by  phenylalanine.

The research was supported by National Institutes of Health (NIH) grant 5R01-NS100081 and a grant from the National PKU Alliance. It was also funded in part through the NIH/National Cancer Institute Cancer Center Support Grant P30 CA006927.

      

The Hospital of Fox Chase Cancer Center and its affiliates (collectively “Fox Chase Cancer Center”), a member of the Temple University Health System, is one of the leading cancer research and treatment centers in the United States. Founded in 1904 in Philadelphia as one of the nation’s first cancer hospitals, Fox Chase was also among the first institutions to be designated a National Cancer Institute Comprehensive Cancer Center in 1974. Fox Chase researchers have won the highest awards in their fields, including two Nobel Prizes. Fox Chase physicians are also routinely recognized in national rankings, and the Center’s nursing program has received the Magnet recognition for excellence five consecutive times. Today, Fox Chase conducts a broad array of nationally competitive basic, translational, and clinical research, with special programs in cancer prevention, detection, survivorship and community outreach. It is the policy of Fox Chase Cancer Center that there shall be no exclusion from, or participation in, and no one denied the benefits of, the delivery of quality medical care on the basis of race, ethnicity, religion, sexual orientation, gender, gender identity/expression, disability, age, ancestry, color, national origin, physical ability, level of education, or source of payment.

 

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