PDBsum entry: 1aup

Oxidoreductase

PDB id: 1aup

Contents

Protein chain

427 a.a. *

Waters ×40

* Residue conservation analysis

PDB id:

1aup

Name:

Oxidoreductase

Title:

Glutamate dehydrogenase

Structure:

NAD-specific glutamate dehydrogenase. Chain: a. Engineered: yes. Mutation: yes

Source:

Clostridium symbiosum. Organism_taxid: 1512. Gene: gdh. Expressed in: escherichia coli. Expression_system_taxid: 562.

Biol. unit:

Homo-Hexamer (from PDB file)

Resolution:

2.50Å R-factor: 0.190

Authors:

P.J.Baker,M.L.Waugh,T.J.Stillman,A.P.Turnbull,D.W.Rice

Key ref:

P.J.Baker et al. (1997). Determinants of substrate specificity in the superfamily of amino acid dehydrogenases. Biochemistry, 36, 16109-16115. PubMed id: 9405044 DOI: 10.1021/bi972024x

Date:

01-Sep-97 Release date: 18-Mar-98

Protein chain

P24295  (DHE2_CLOSY) -  NAD-specific glutamate dehydrogenase

Seq: 450 a.a.

Struc: 427 a.a.*

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

Enzyme reactions

• Enzyme class: E.C.1.4.1.2 - Glutamate dehydrogenase.
• Reaction: L-glutamate + H₂O + NAD⁺ = 2-oxoglutarate + NH₃ + NADH

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

 Gene Ontology (GO) functional annotation 

• Biological process: oxidation-reduction process (2 terms)
• Biochemical function: nucleotide binding (4 terms)

reference

DOI no: 10.1021/bi972024x

Biochemistry 36:16109-16115 (1997)

PubMed id: 9405044

Determinants of substrate specificity in the superfamily of amino acid dehydrogenases.

P.J.Baker, M.L.Waugh, X.G.Wang, T.J.Stillman, A.P.Turnbull, P.C.Engel, D.W.Rice.

ABSTRACT

The subunit of the enzyme glutamate dehydrogenase comprises two domains separated by a cleft harboring the active site. One domain is responsible for dinucleotide binding and the other carries the majority of residues which bind the substrate. During the catalytic cycle a large movement between the two domains occurs, closing the cleft and bringing the C4 of the nicotinamide ring and the Calpha of the substrate into the correct positioning for hydride transfer. In the active site, two residues, K89 and S380, make interactions with the gamma-carboxyl group of the glutamate substrate. In leucine dehydrogenase, an enzyme belonging to the same superfamily, the equivalent residues are L40 and V294, which create a more hydrophobic specificity pocket and provide an explanation for their differential substrate specificity. In an attempt to change the substrate specificity of glutamate dehydrogenase toward that of leucine dehydrogenase, a double mutant, K89L,S380V, of glutamate dehydrogenase has been constructed. Far from having a high specificity for leucine, this mutant appears to be devoid of any catalytic activity over a wide range of substrates tested. Determination of the three-dimensional structure of the mutant enzyme has shown that the loss of function is related to a disordering of residues linking the enzyme's two domains, probably arising from a steric clash between the valine side chain, introduced at position 380 in the mutant, and a conserved threonine residue, T193. In leucine dehydrogenase the steric clash between the equivalent valine and threonine side chains (V294, T134) does not occur owing to shifts of the main chain to which these side chains are attached. Thus, the differential substrate specificity seen in the amino acid dehydrogenase superfamily arises from both the introduction of simple point mutations and the fine tuning of the active site pocket defined by small but significant main chain rearrangements.