Redox-switch modulation of human SSADH by dynamic catalytic loop

Abstract

Succinic semialdehyde dehydrogenase (SSADH) is involved in the final degradation step of the inhibitory neurotransmitter c-aminobutyric acid by converting succinic semialdehyde to succinic acid in the mitochondrial matrix. SSADH deficiency, a rare autosomal recessive disease, exhibits variable clinical phenotypes, including psychomotor retardation, language delay, behaviour disturbance and convulsions. Here, we present crystal structures of both the oxidized and reduced forms of human SSADH. Interestingly, the structures show that the catalytic loop of the enzyme undergoes large structural changes depending on the redox status of the environment, which is mediated by a reversible disulphide bond formation between a catalytic Cys340 and an adjacent Cys342 residues located on the loop. Subsequent in vivo and in vitro studies reveal that the 'dynamic catalytic loop' confers a response to reactive oxygen species and changes in redox status, indicating that the redox-switch modulation could be a physiological control mechanism of human SSADH. Structural basis for the substrate specificity of the enzyme and the impact of known missense point mutations associated with the disease pathogenesis are presented as well.

Structural Basis for a Cofactor-dependent Oxidation Protection and Catalysis of Cyanobacterial Succinic Semialdehyde Dehydrogenase

“…The overall structure of monomeric SySSADH is reminiscent of an ALDH fold (22) and similar to the structures of HsSSADH (11) and EcSSADH (12,13). It is made up of three segments: ␣/␤-fold N-and C-domains for a cofactor binding and a catalytic domain, respectively, and three antiparallel ␤-strands constituting a dimerization domain (Fig.…”

“…the second cysteine in the catalytic loop, and that the reduced cysteine is required for activity (11). Due to the lack of a second cysteine, such a mechanism is unavailable in SySSADH as well as in other members of the 1-Cys SSADH family.…”

“…1C), which form a reversible disulfide bridge in response to redox conditions. As a result, the loop undergoes a large conformational transition between a functionally active open form in the reduced state and a catalytically inactive, closed form in the oxidized state (11). In contrast, the structural features of 2-Cys E. coli SSADH (EcSSADH) were investigated only in its reduced form (12,13).…”

“…SSADH belongs to the aldehyde dehydrogenase (ALDH) superfamily (9,10), and its structure in humans (11) and Escherichia coli (12,13) has only recently become available. These investigations highlighted a redox-dependent regulation of SSADH activity.…”

Methylmalonate-semialdehyde Dehydrogenase from Bacillus subtilis

“…From an evolutionary point of view, it is interesting to note that such a stabilization mode is not unique within the ALDH superfamily. Indeed, in the succinate semialdehyde dehydrogenase, a member of the hydrolytic ALDH family, the stabilization of the carboxylate group of the substrate is mainly achieved through hydrogen bonds with two arginine (Arg-213 and Arg-334) residues (29). Although differing in position in primary structures, superimposition of the GAPN, MSDH, and succinate semialdehyde dehydrogenase active sites shows that Arg-213 and Arg-334 are structurally and functionally equivalents to Arg-124 and Arg-301, respectively (not shown).…”