Abstract
BackgroundIn mammals succinic semialdehyde dehydrogenase (SSADH) plays an essential role in the metabolism of the inhibitory neurotransmitter γ-aminobutyric acid (GABA) to succinic acid (SA). Deficiency of SSADH in humans results in elevated levels of GABA and γ-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 FindingsHere 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/SignificanceOur 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.
Highlights
Succinic semialdehyde dehydrogenase (SSADH) belongs to the aldehyde dehydrogenases (ALDH) superfamily [1] and has been identified and purified from mammals [2,3,4,5] as well as from microorganisms [6,7,8]
Production and Characterisation of E. coli succinic semialdehyde dehydrogenase (SSADH) Recombinant E. coli SSADH was purified as a tetrameric molecule which is in agreement with the previous description in the literature [6]
The enzyme activity measured in the presence of NADP+ is approximately 20-fold higher than that measured in the presence of NAD+ as described previously [27]
Summary
Succinic semialdehyde dehydrogenase (SSADH) belongs to the aldehyde dehydrogenases (ALDH) superfamily [1] and has been identified and purified from mammals [2,3,4,5] as well as from microorganisms [6,7,8]. GABA-transaminase (EC 2.6.1.19) catalyses the breakdown of GABA in the presence of a-ketoglutarate to produce succinic semialdehyde (SSA) and glutamic acid (Figure 1). SSA is converted to succinic acid (SA) by the NAD+/NADP+-dependant enzyme succinic semialdehyde dehydrogenase (SSADH; EC 1.2.1.24) [11]. SSA can be converted to c-hydroxybutyric acid (GHB) by succinic semialdehyde reductase [12] (see Figure 1). In mammals succinic semialdehyde dehydrogenase (SSADH) plays an essential role in the metabolism of the inhibitory neurotransmitter c-aminobutyric acid (GABA) to succinic acid (SA). Deficiency of SSADH in humans results in elevated levels of GABA and c-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
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