PDBsum entry: 1il0

Oxidoreductase

PDB id: 1il0

Contents

Protein chains

291 a.a. *

Ligands

CAA ×2

NAD ×2

Waters ×242

* Residue conservation analysis

PDB id:

1il0

Name:

Oxidoreductase

Title:

X-ray crystal structure of the e170q mutant of human l-3- hydroxyacyl-coa dehydrogenase

Structure:

3-hydroxyacyl-coa dehydrogenase. Chain: a, b. Synonym: hcdh. Engineered: yes. Mutation: yes

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: hcdh. Expressed in: escherichia coli. Expression_system_taxid: 562.

Biol. unit:

Dimer (from PQS)

Resolution:

2.20Å R-factor: 0.219 R-free: 0.276

Authors:

J.J.Barycki,L.K.O'Brien,A.W.Strauss,L.J.Banaszak

Key ref:

J.J.Barycki et al. (2001). Glutamate 170 of human l-3-hydroxyacyl-CoA dehydrogenase is required for proper orientation of the catalytic histidine and structural integrity of the enzyme. J Biol Chem, 276, 36718-36726. PubMed id: 11451959 DOI: 10.1074/jbc.M104839200

Date:

07-May-01 Release date: 07-Nov-01

Protein chains

Q16836  (HCDH_HUMAN) -  Hydroxyacyl-coenzyme A dehydrogenase, mitochondrial

Seq: 314 a.a.

Struc: 291 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.1.1.1.35 - 3-hydroxyacyl-CoA dehydrogenase.
• Reaction: (S)-3-hydroxyacyl-CoA + NAD⁺ = 3-oxoacyl-CoA + NADH

(S)-3-hydroxyacyl-CoA (Bound ligand (Het Group name = CAA) matches with 96.00% similarity) + NAD(+) (Bound ligand (Het Group name = NAD) corresponds exactly) = 3-oxoacyl-CoA + NADH

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

 Gene Ontology (GO) functional annotation 

• Cellular component: cytoplasm (4 terms)
• Biological process: response to activity (10 terms)
• Biochemical function: nucleotide binding (6 terms)

reference

DOI no: 10.1074/jbc.M104839200

J Biol Chem 276:36718-36726 (2001)

PubMed id: 11451959

Glutamate 170 of human l-3-hydroxyacyl-CoA dehydrogenase is required for proper orientation of the catalytic histidine and structural integrity of the enzyme.

J.J.Barycki, L.K.O'Brien, A.W.Strauss, L.J.Banaszak.

ABSTRACT

l-3-Hydroxyacyl-CoA dehydrogenase (HAD), the penultimate enzyme in the beta-oxidation spiral, reversibly catalyzes the conversion of l-3-hydroxyacyl-CoA to the corresponding 3-ketoacyl-CoA. Similar to other dehydrogenases, HAD contains a general acid/base, His(158), which is within hydrogen bond distance of a carboxylate, Glu(170). To investigate its function in this catalytic dyad, Glu(170) was replaced with glutamine (E170Q), and the mutant enzyme was characterized. Whereas substrate and cofactor binding were unaffected by the mutation, E170Q exhibited diminished catalytic activity. Protonation of the catalytic histidine did not restore wild-type activity, indicating that modulation of the pK(a) of His(158) is not the sole function of Glu(170). The pH profile of charge transfer complex formation, an independent indicator of active site integrity, was unaltered by the amino acid substitution, but the intensity of the charge transfer band was diminished. This observation, coupled with significantly reduced enzymatic stability of the E170Q mutant, implicates Glu(170) in maintenance of active site architecture. Examination of the crystal structure of E170Q in complex with NAD(+) and acetoacetyl-CoA (R = 21.9%, R(free) = 27.6%, 2.2 A) reveals that Gln(170) no longer hydrogen bonds to the side chain of His(158). Instead, the imidazole ring is nearly perpendicular to its placement in the comparable native complex and no longer positioned for efficient catalysis.

Selected figure(s)

Figure 4.
Fig. 4. 2|F[o]| |F[c]| simulated annealing omit map of the E170Q mutant active site. Gln170, His158, NAD^+, and AACoA as well as residues and water molecules within 3.5 Å of these groups were omitted, and the remaining model was subjected to simulated annealing prior to map calculation. Shown are the resultant map contoured at 1.25 , the final E170Q model (dark gray), and, for reference, the corresponding native structure (light gray). The electron density map supports an E170Q model in which the imidazole ring of His158 has rotated ~90° relative to its placement in the native HAD structure, while adjacent residues are essentially undisturbed. Relatively weak electron density, which has been omitted for clarity, is observed for the structural water molecule of the E170Q model, Wat 984.

Figure 5.
Fig. 5. Comparison of the enzyme active sites of native and E170Q L-3-hydroxyacyl-CoA dehydrogenases. The stereo diagrams depict the enzyme active sites of native HAD ( A) and E170Q HAD (B) in complex with NAD^+ and AACoA. Relevant interatomic distances within 3.5 Å are illustrated as dotted black lines, with oxygen atoms colored in red, nitrogen in blue, carbon in green, sulfur in yellow, and phosphorus in purple. The imidazole ring of His158 adopts a significantly different conformation in the E170Q mutant structure as compared with the native model. Additional differences are discussed under "Results."

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2001, 276, 36718-36726) copyright 2001.

Figures were selected by an automated process.