Crystal Structures of Wild-Type and F448A Mutant Citrobacter freundii Tyrosine Phenol-Lyase Complexed with a Substrate and Inhibitors: Implications for the Reaction Mechanism

Journal Article · Thu Sep 27 00:00:00 EDT 2018 · Biochemistry

DOI:https://doi.org/10.1021/acs.biochem.8b00724· OSTI ID:1483076

Phillips, Robert S. ^[1]; Craig, Steven ^[1]

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1. Univ. of Georgia, Athens, GA (United States)

Tyrosine phenol-lyase (TPL; EC 4.1.99.2) is a pyridoxal 5'-phosphate-dependent enzyme that catalyzes the reversible hydrolytic cleavage of l-tyrosine to phenol and ammonium pyruvate. We have shown previously that F448A TPL has k_cat and k_cat/K_m values for l-tyrosine reduced by ~10⁴-fold. We have now obtained crystal structures of F448A TPL and complexes with l-alanine, l-methionine, l-phenylalanine, and 3-F-l-tyrosine at 2.05–2.27 Å and the complex of wild-type TPL with l-phenylalanine at 1.8 Å. The small domain of F448A TPL, where Phe-448 is located, is more disordered in chain A than in wild-type TPL. The complexes of F448A TPL with l-alanine and l-phenylalanine are in an open conformation in both chains, while the complex with l-methionine is a 52:48 open:closed equilibrium mixture in chain A. Wild-type TPL with l-alanine is closed in chain A and open in chain B, and the complex with l-phenylalanine is a 56:44 open:closed mixture in chain A. Thus, the Phe-448 to alanine mutation affects the conformational equilibrium of open and closed active sites. The structure of the 3-F-l-tyrosine quinonoid complex of F448A TPL is unstrained and in an open conformation, with a hydrogen bond from the phenolic OH to Thr-124. Furthermore, these results support our previous conclusion that ground-state strain plays a critical role in the mechanism of TPL.

• Research Organization: Argonne National Laboratory (ANL), Argonne, IL (United States)
• Sponsoring Organization: National Inst. of Health; USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22)
• Grant/Contract Number: W-31-109-ENG-3
• OSTI ID: 1483076
• Journal Information: Biochemistry, Journal Name: Biochemistry Journal Issue: 43 Vol. 57; ISSN 0006-2960
• Publisher: American Chemical Society (ACS)
• Country of Publication: United States
• Language: ENGLISH

Related Subjects

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY
59 BASIC BIOLOGICAL SCIENCES
aromatic compounds
chemical structure
conformation
noncovalent interactions
peptides and proteins