Solved Protein Structure Holds Answers to Metabolic Disease That Afflicts Infants

This shows the first X-ray crystal structure of phenylalanine hydroxylase (PAH), the enzyme that is defective in patients with the metabolic disorder PKU. The structure was recently determined by researchers and published in the Proceedings of the National Academy of Sciences in February.

Drexel University researchers and an international group of scientists have made a critical step toward understanding a disorder that affects patients from birth into adulthood.

Phenylketonuria, or PKU, is an inherited metabolic disease that affects one in 20,000 babies born in the United States each year. The enzyme that breaks down the amino acid phenylalanine is either missing or defective in newborns with the disorder, so patients must adhere to a protein-restricted diet throughout their lives to prevent mental and behavioral abnormalities.

But maintaining this strict diet can be a challenge, which intensifies the need for new, non-dietary treatments to combat PKU symptoms.

In a study recently published in Proceedings of the National Academy of Sciences, researchers made significant strides toward that goal by determining the structure of phenylalanine hydroxylase (PAH) — the enzyme that is defective in PKU patients.

“We have solved the first X-ray crystal structure of full-length PAH, which has defied crystal structure determination for decades,” said the study’s principal investigator Eileen K. Jaffe, professor of Molecular Therapeutics at Fox Chase Cancer Center – Temple Health. “This structure will help us understand the molecular origins of PKU and is an important advance in developing much-needed drugs for patients.”

Defects in the PAH enzyme cause phenylalanine to build up to toxic levels in the body. Because nerve cells in the brain are particularly sensitive to phenylalanine levels, excessive amounts of this substance can cause brain defects.

While researchers have tried for decades to determine the structure of the PAH enzyme, they have been unsuccessful, said Emilia Arturo, a graduate student in Drexel’s College of Medicine and study co-author.

“Knowing the structure at this high resolution is expected to speed up any current drug discovery pursuits that would otherwise be based only on models of the full-length protein,” Arturo said.

The research team purified the enzyme and then crystalized the proteins, before they were analyzed with X-ray machines and illustrated into a 3-D image. Arturo and Patrick Loll, a professor in the College of Medicine’s Department of Biochemistry and Molecular Biology, were responsible for data acquisition and structuring the solution.

The researchers ultimately discovered that the PAH enzyme is represented by two distinct structures rather than one. Once the researchers could separate the two, they were able to grow crystals where scientists have not been able to in the past.  

Using what they have learned from this study, the team is now working on solving the structure of activated PAH, which functions to prevent blood phenylalanine from rising to neurotoxic levels. 

“Knowledge of both PAH structures will help us use structure-based drug design techniques to develop drugs that will favor activated PAH as a therapeutic approach,” Jaffe said.

In the end, this research could pave the way for new treatment strategies not only for PKU, but also for cancer and other diseases.

“Both inborn errors of metabolism and cancer occur when our proteins do not function as they should,” Jaffe said. “Our focus on how different protein assemblies can be harnessed to control protein function has tremendous potential for drug discovery. However, this approach requires that protein chemists and pharmaceutical companies embrace an expanded view of how small molecule therapeutics can work.”

The international research team included contributors from Drexel University, Fox Chase Cancer Center – Temple Health, the University of Pennsylvania Perelman School of Medicine and the University of Canterbury in New Zealand.