The liver enzyme phenylalanine hydroxylase is responsible for catabolism of excess phenylalanine in the diet. Deficiencies in levels of the enzyme result in the metabolic disorder phenylketonuria, a disease with devastating neurological consequences if untreated, demonstrating the physiological importance of the enzyme. Central to the proper function of phenylalanine hydroxylase is the regulation of the enzyme by its substrates, phenylalanine and tetrahydrobiopterin, and by phosphorylation. Both forms of regulation require the N-terminal ~117 residue regulatory domain. In the generally-accepted model for regulation, the resting form of the enzyme is inactive and allosteric binding of phenylalanine at a regulatory site converts the enzyme to an active form. Tetrahydrobiopterin stabilizes the inactive form, while phosphorylation potentiates the conversion to the active form. The goal of the research proposed here is to understand the structural and dynamic basis for the allosteric regulation. The proposed experiments will combine modern structural approaches (e.g., NMR and mass spectroscopy) with measurement of intrinsic rate constants for binding and catalysis in order to provide a more complete understanding of the allosteric regulation of phenylalanine hydroxylase.

Public Health Relevance

This is a proposal to study the regulation of the liver enzyme phenylalanine hydroxylase. An inherited deficiency in this enzyme unless the affected individual is placed on a restricted diet from birth. A better understanding of the regulation will improve our ability to develop better treatments for PKU.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM098140-04
Application #
8663288
Study Section
Special Emphasis Panel ()
Program Officer
Anderson, Vernon
Project Start
2011-09-20
Project End
2015-05-31
Budget Start
2014-06-01
Budget End
2015-05-31
Support Year
4
Fiscal Year
2014
Total Cost
$336,010
Indirect Cost
$109,197
Name
University of Texas Health Science Center San Antonio
Department
Biochemistry
Type
Schools of Medicine
DUNS #
800772162
City
San Antonio
State
TX
Country
United States
Zip Code
78229
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Zhang, Shengnan; Huang, Tao; Ilangovan, Udayar et al. (2014) The solution structure of the regulatory domain of tyrosine hydroxylase. J Mol Biol 426:1483-97
Roberts, Kenneth M; Pavon, Jorge Alex; Fitzpatrick, Paul F (2013) Kinetic mechanism of phenylalanine hydroxylase: intrinsic binding and rate constants from single-turnover experiments. Biochemistry 52:1062-73
Daubner, S Colette; Avila, Audrey; Bailey, Johnathan O et al. (2013) Mutagenesis of a specificity-determining residue in tyrosine hydroxylase establishes that the enzyme is a robust phenylalanine hydroxylase but a fragile tyrosine hydroxylase. Biochemistry 52:1446-55
Li, Jun; Fitzpatrick, Paul F (2013) Regulation of phenylalanine hydroxylase: conformational changes upon phosphorylation detected by H/D exchange and mass spectrometry. Arch Biochem Biophys 535:115-9
Fitzpatrick, Paul F (2012) Allosteric regulation of phenylalanine hydroxylase. Arch Biochem Biophys 519:194-201