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The X-ray crystal structure of Escherichia coli succinic semialdehyde dehydrogenase; structural insights into NADP+/enzyme interactions.

Langendorf CG, Key TL, Fenalti G, Kan WT, Buckle AM, Caradoc-Davies T, Tuck KL, Law RH, Whisstock JC - PLoS ONE (2010)

Bottom Line: In the E. coli SSADH structure, electron density for the complete NADP+ cofactor in the binding sites is clearly evident; these data in particular revealing how the nicotinamide ring of the cofactor is positioned in each active site.Our structural data suggest that a deletion of three amino acids in E. coli SSADH permits this enzyme to use NADP+, whereas in contrast the human enzyme utilises NAD+.Furthermore, the structure of E. coli SSADH gives additional insight into human mutations that result in disease.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Monash University, Clayton Campus, Melbourne, Victoria, Australia.

ABSTRACT

Background: In mammals succinic semialdehyde dehydrogenase (SSADH) plays an essential role in the metabolism of the inhibitory neurotransmitter gamma-aminobutyric acid (GABA) to succinic acid (SA). Deficiency of SSADH in humans results in elevated levels of GABA and gamma-Hydroxybutyric acid (GHB), which leads to psychomotor retardation, muscular hypotonia, non-progressive ataxia and seizures. In Escherichia coli, two genetically distinct forms of SSADHs had been described that are essential for preventing accumulation of toxic levels of succinic semialdehyde (SSA) in cells.

Methodology/principal findings: Here we structurally characterise SSADH encoded by the E coli gabD gene by X-ray crystallographic studies and compare these data with the structure of human SSADH. In the E. coli SSADH structure, electron density for the complete NADP+ cofactor in the binding sites is clearly evident; these data in particular revealing how the nicotinamide ring of the cofactor is positioned in each active site.

Conclusions/significance: Our structural data suggest that a deletion of three amino acids in E. coli SSADH permits this enzyme to use NADP+, whereas in contrast the human enzyme utilises NAD+. Furthermore, the structure of E. coli SSADH gives additional insight into human mutations that result in disease.

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Related in: MedlinePlus

Different binding states of NAD(P)+ in the ALDH family.Four different conformations of NAD(P)+ are shown as sticks, hydride conformation (PDB ID: 1bpw[31]: yellow), hydrolysis conformation (E. coli SSADH: green), out conformation (PDB ID: 2ilu [29]: magenta) and flexible, where the nicotinamide ribose moiety is unable to be resolved using X-ray crystallography (PDB ID: 2w8r [26]: blue). The general base (E254) and the catalytic cysteine (C288: both orange), which are conserved in human and E. coli SSADH and the whole ALDH family, have been labelled to define the active site.
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pone-0009280-g007: Different binding states of NAD(P)+ in the ALDH family.Four different conformations of NAD(P)+ are shown as sticks, hydride conformation (PDB ID: 1bpw[31]: yellow), hydrolysis conformation (E. coli SSADH: green), out conformation (PDB ID: 2ilu [29]: magenta) and flexible, where the nicotinamide ribose moiety is unable to be resolved using X-ray crystallography (PDB ID: 2w8r [26]: blue). The general base (E254) and the catalytic cysteine (C288: both orange), which are conserved in human and E. coli SSADH and the whole ALDH family, have been labelled to define the active site.

Mentions: Amongst the well defined cofactor crystal structures, three different conformations of NAD(P)+ found in the ALDH superfamily have been described [29]–the hydride transfer (PDB ID: 1bpw [31]), the hydrolysis (see below) and the “out” conformation (PDB ID: 2ilu [29]; Figure 7). Superposition with ALDH structures in the hydrolysis conformation (PDB ID: 1bxs and 1O01 [34], [35]) reveals that the cofactor in the E. coli SSADH structure most closely resembles this conformation, where the nicotinamide ring is retracted from the active site such that the general base E251 is now situated in an ideal distance (3.69 Å to the substrate SSA and 3.15 Å to the catalytic cysteine C288) to catalyse the deacylation process.


The X-ray crystal structure of Escherichia coli succinic semialdehyde dehydrogenase; structural insights into NADP+/enzyme interactions.

Langendorf CG, Key TL, Fenalti G, Kan WT, Buckle AM, Caradoc-Davies T, Tuck KL, Law RH, Whisstock JC - PLoS ONE (2010)

Different binding states of NAD(P)+ in the ALDH family.Four different conformations of NAD(P)+ are shown as sticks, hydride conformation (PDB ID: 1bpw[31]: yellow), hydrolysis conformation (E. coli SSADH: green), out conformation (PDB ID: 2ilu [29]: magenta) and flexible, where the nicotinamide ribose moiety is unable to be resolved using X-ray crystallography (PDB ID: 2w8r [26]: blue). The general base (E254) and the catalytic cysteine (C288: both orange), which are conserved in human and E. coli SSADH and the whole ALDH family, have been labelled to define the active site.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2823781&req=5

pone-0009280-g007: Different binding states of NAD(P)+ in the ALDH family.Four different conformations of NAD(P)+ are shown as sticks, hydride conformation (PDB ID: 1bpw[31]: yellow), hydrolysis conformation (E. coli SSADH: green), out conformation (PDB ID: 2ilu [29]: magenta) and flexible, where the nicotinamide ribose moiety is unable to be resolved using X-ray crystallography (PDB ID: 2w8r [26]: blue). The general base (E254) and the catalytic cysteine (C288: both orange), which are conserved in human and E. coli SSADH and the whole ALDH family, have been labelled to define the active site.
Mentions: Amongst the well defined cofactor crystal structures, three different conformations of NAD(P)+ found in the ALDH superfamily have been described [29]–the hydride transfer (PDB ID: 1bpw [31]), the hydrolysis (see below) and the “out” conformation (PDB ID: 2ilu [29]; Figure 7). Superposition with ALDH structures in the hydrolysis conformation (PDB ID: 1bxs and 1O01 [34], [35]) reveals that the cofactor in the E. coli SSADH structure most closely resembles this conformation, where the nicotinamide ring is retracted from the active site such that the general base E251 is now situated in an ideal distance (3.69 Å to the substrate SSA and 3.15 Å to the catalytic cysteine C288) to catalyse the deacylation process.

Bottom Line: In the E. coli SSADH structure, electron density for the complete NADP+ cofactor in the binding sites is clearly evident; these data in particular revealing how the nicotinamide ring of the cofactor is positioned in each active site.Our structural data suggest that a deletion of three amino acids in E. coli SSADH permits this enzyme to use NADP+, whereas in contrast the human enzyme utilises NAD+.Furthermore, the structure of E. coli SSADH gives additional insight into human mutations that result in disease.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Monash University, Clayton Campus, Melbourne, Victoria, Australia.

ABSTRACT

Background: In mammals succinic semialdehyde dehydrogenase (SSADH) plays an essential role in the metabolism of the inhibitory neurotransmitter gamma-aminobutyric acid (GABA) to succinic acid (SA). Deficiency of SSADH in humans results in elevated levels of GABA and gamma-Hydroxybutyric acid (GHB), which leads to psychomotor retardation, muscular hypotonia, non-progressive ataxia and seizures. In Escherichia coli, two genetically distinct forms of SSADHs had been described that are essential for preventing accumulation of toxic levels of succinic semialdehyde (SSA) in cells.

Methodology/principal findings: Here we structurally characterise SSADH encoded by the E coli gabD gene by X-ray crystallographic studies and compare these data with the structure of human SSADH. In the E. coli SSADH structure, electron density for the complete NADP+ cofactor in the binding sites is clearly evident; these data in particular revealing how the nicotinamide ring of the cofactor is positioned in each active site.

Conclusions/significance: Our structural data suggest that a deletion of three amino acids in E. coli SSADH permits this enzyme to use NADP+, whereas in contrast the human enzyme utilises NAD+. Furthermore, the structure of E. coli SSADH gives additional insight into human mutations that result in disease.

Show MeSH
Related in: MedlinePlus