PDBsum entry 1c1x

Oxidoreductase

PDB id: 1c1x

Contents

Protein chains

348 a.a. *

Ligands

HFA ×2

NAD ×2

PO4

IPA ×3

Metals

__K ×4

_NA

Waters ×844

* Residue conservation analysis

PDB id:

1c1x

Name:

Oxidoreductase

Title:

L-phenylalanine dehydrogenase structure in ternary complex with NAD+ and l-3-phenyllactate

Structure:

L-phenylalanine dehydrogenase. Chain: a. Engineered: yes. Protein (l-phenylalanine dehydrogenase). Chain: b. Engineered: yes

Source:

Rhodococcus sp.. Organism_taxid: 1831. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008. Organism_taxid: 1831

Resolution:

1.40Å R-factor: 0.183 R-free: 0.229

Authors:

J.L.Vanhooke,J.B.Thoden

Key ref:

N.M.Brunhuber et al. (2000). Rhodococcus L-phenylalanine dehydrogenase: kinetics, mechanism, and structural basis for catalytic specificity. Biochemistry, 39, 9174-9187. PubMed id: 10924111 DOI: 10.1021/bi000494c

Date:

22-Jul-99 Release date: 30-Aug-00

Protein chains

Q59771  (DHPH_RHOSO) -  Phenylalanine dehydrogenase from Rhodococcus sp

Seq: 356 a.a.

Struc: 348 a.a.*

* PDB and UniProt seqs differ at 3 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.1.4.1.20 - phenylalanine dehydrogenase.
• Reaction: L-phenylalanine + NAD⁺ + H2O = 3-phenylpyruvate + NH4⁺ + NADH + H⁺

L-phenylalanine + NAD(+) (Bound ligand (Het Group name = HFA) matches with 84.62% similarity) + H2O (Bound ligand (Het Group name = NAD) corresponds exactly) = 3-phenylpyruvate + NH4(+) + NADH + H(+)

• Cofactor: Ca(2+)

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1021/bi000494c

Biochemistry 39:9174-9187 (2000)

PubMed id: 10924111

Rhodococcus L-phenylalanine dehydrogenase: kinetics, mechanism, and structural basis for catalytic specificity.

N.M.Brunhuber, J.B.Thoden, J.S.Blanchard, J.L.Vanhooke.

ABSTRACT

Phenylalanine dehydrogenase catalyzes the reversible, pyridine nucleotide-dependent oxidative deamination of L-phenylalanine to form phenylpyruvate and ammonia. We have characterized the steady-state kinetic behavior of the enzyme from Rhodococcus sp. M4 and determined the X-ray crystal structures of the recombinant enzyme in the complexes, E.NADH.L-phenylalanine and E.NAD(+). L-3-phenyllactate, to 1.25 and 1.4 A resolution, respectively. Initial velocity, product inhibition, and dead-end inhibition studies indicate the kinetic mechanism is ordered, with NAD(+) binding prior to phenylalanine and the products' being released in the order of ammonia, phenylpyruvate, and NADH. The enzyme shows no activity with NADPH or other 2'-phosphorylated pyridine nucleotides but has broad activity with NADH analogues. Our initial structural analyses of the E.NAD(+).phenylpyruvate and E.NAD(+). 3-phenylpropionate complexes established that Lys78 and Asp118 function as the catalytic residues in the active site [Vanhooke et al. (1999) Biochemistry 38, 2326-2339]. We have studied the ionization behavior of these residues in steady-state turnover and use these findings in conjunction with the structural data described both here and in our first report to modify our previously proposed mechanism for the enzymatic reaction. The structural characterizations also illuminate the mechanism of the redox specificity that precludes alpha-amino acid dehydrogenases from functioning as alpha-hydroxy acid dehydrogenases.