Title: First structure of full-length mammalian phenylalanine hydroxylase reveals the architecture of an autoinhibited tetramer

Abstract

Improved understanding of the relationship among structure, dynamics, and function for the enzyme phenylalanine hydroxylase (PAH) can lead to needed new therapies for phenylketonuria, the most common inborn error of amino acid metabolism. PAH is a multidomain homo-multimeric protein whose conformation and multimerization properties respond to allosteric activation by the substrate phenylalanine (Phe); the allosteric regulation is necessary to maintain Phe below neurotoxic levels. A recently introduced model for allosteric regulation of PAH involves major domain motions and architecturally distinct PAH tetramers [Jaffe EK, Stith L, Lawrence SH, Andrake M, Dunbrack RL, Jr (2013) Arch Biochem Biophys 530(2):73–82]. Herein, we present, to our knowledge, the first X-ray crystal structure for a full-length mammalian (rat) PAH in an autoinhibited conformation. Chromatographic isolation of a monodisperse tetrameric PAH, in the absence of Phe, facilitated determination of the 2.9 Å crystal structure. The structure of full-length PAH supersedes a composite homology model that had been used extensively to rationalize phenylketonuria genotype–phenotype relationships. Small-angle X-ray scattering (SAXS) confirms that this tetramer, which dominates in the absence of Phe, is different from a Phe-stabilized allosterically activated PAH tetramer. The lack of structural detail for activated PAH remains a barrier to complete understanding of phenylketonuria genotype–phenotypemore » relationships. Nevertheless, the use of SAXS and X-ray crystallography together to inspect PAH structure provides, to our knowledge, the first complete view of the enzyme in a tetrameric form that was not possible with prior partial crystal structures, and facilitates interpretation of a wealth of biochemical and structural data that was hitherto impossible to evaluate.« less

Authors:

Arturo, Emilia C.; Gupta, Kushol; Héroux, Annie; Stith, Linda; Cross, Penelope J.; Parker, Emily J.; Loll, Patrick J.; Jaffe, Eileen K.

Publication Date:

2016-02-16

Research Org.:

Brookhaven National Lab. (BNL), Upton, NY (United States)

Sponsoring Org.:

USDOE Office of Science (SC), Basic Energy Sciences (BES); USDOE Office of Science (SC), Biological and Environmental Research (BER); National Cancer Institute Comprehensive Cancer Center; National Center for Research Resources of the National Inst. of Health

OSTI Identifier:

1238204

Alternate Identifier(s):

OSTI ID: 1354369

Report Number(s):

BNL-112885-2016-JA
Journal ID: ISSN 0027-8424; /pnas/113/9/2394.atom

Grant/Contract Number:  

AC02-98CH10886; SC00112704; P41GM103473; P41RR012408; P30CA006927

Resource Type:

Published Article

Journal Name:

Proceedings of the National Academy of Sciences of the United States of America

Additional Journal Information:

Journal Name: Proceedings of the National Academy of Sciences of the United States of America Journal Volume: 113 Journal Issue: 9; Journal ID: ISSN 0027-8424

Publisher:

National Academy of Sciences

Country of Publication:

United States

Language:

English

Subject:

59 BASIC BIOLOGICAL SCIENCES; phenylalanine hydroxylase; phenylketonuria; X-ray crystallography; small-angle X-ray scattering; allosteric regulation