Adenine Binding Mode Is a Key Factor in Triggering the Early Release of NADH in Coenzyme A-dependent Methylmalonate Semialdehyde Dehydrogenase

Bchini, Raphaël; Dubourg-Gerecke, Hélène; Rahuel-Clermont, Sophie; Aubry, A.; Branlant, Guy; Didierjean, Claude; Talfournier, François

doi:10.1074/jbc.m112.350272

Cited by 5 publications

(6 citation statements)

References 39 publications

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“…A disorder to order transition has been proposed to be linked to catalysis and to act as a mechanism conferring substrate specificity for retinal vs. short chain aldehydes [23, 24]. This hypothesis is consistent with the general notion that in ALDH enzymes, structural flexibility within the active site induced by ligand binding or associated with cofactor dynamics is an integral component of catalysis [25–32]. …”

Section: Introductionsupporting

confidence: 62%

Retinoic acid biosynthesis catalyzed by retinal dehydrogenases relies on a rate-limiting conformational transition associated with substrate recognition

Bchini¹,

Vasiliou²,

Branlant³

et al. 2013

Chemico-Biological Interactions

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Retinoic acid (RA), a metabolite of vitamin A, exerts pleiotropic effects throughout life in vertebrate organisms. Thus, RA action must be tightly regulated through the coordinated action of biosynthetic and degradating enzymes. The last step of retinoic acid biosynthesis is irreversibly catalyzed by the NAD-dependent retinal dehydrogenases (RALDH), which are members of the aldehyde dehydrogenase (ALDH) superfamily. Low intracellular retinal concentrations imply efficient substrate molecular recognition to ensure high affinity and specificity of RALDHs for retinal. This study addresses the molecular basis of retinal recognition in human ALDH1A1 (or RALDH1) and rat ALDH1A2 (or RALDH2), through the comparison of the catalytic behavior of retinal analogs and use of the fluorescence properties of retinol. We show that, in contrast to long chain unsaturated substrates, the rate-limiting step of retinal oxidation by RALDHs is associated with acylation. Use of the fluorescence resonance energy transfer upon retinol interaction with RALDHs provides evidence that retinal recognition occurs in two steps: binding into the substrate access channel, and a slower structural reorganization with a rate constant of the same magnitude as the kcat for retinal oxidation: 0.18 vs. 0.07 s−1 and 0.25 vs. 0.1 s−1 for ALDH1A1 and ALDH1A2, respectively. This suggests that the conformational transition of the RALDH-retinal complex significantly contributes to the rate-limiting step that controls the kinetics of retinal oxidation, as a prerequisite for the formation of a catalytically competent Michaelis complex. This conclusion is consistent with the general notion that structural flexibility within the active site of ALDH enzymes has been shown to be an integral component of catalysis.

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Section: Introductionsupporting

confidence: 62%

Retinoic acid biosynthesis catalyzed by retinal dehydrogenases relies on a rate-limiting conformational transition associated with substrate recognition

Bchini¹,

Vasiliou²,

Branlant³

et al. 2013

Chemico-Biological Interactions

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“…In hydrolytic ALDHs, this makes the hydrolysis possible by enabling the Glu268 to activate the water molecule, while in their CoA-dependent counterparts it possibly contributes to make the thioacylenzyme competent for transthioesterification. In MSDH, weaker stabilization of the adenine ring triggers early NADH release 24 followed by the β-decarboxylation process, 20 subsequent CoA nucleophilic attack and product release. Except for the decarboxylation, this scenario may hold for all CoA-dependent ALDHs.…”

Section: ■ Discussionmentioning

confidence: 99%

“…Except for the decarboxylation, this scenario may hold for all CoA-dependent ALDHs. In hydrolytic ALDHs, cofactor stabilization mainly results from interactions with adenine moiety, 24 thus explaining the late release of NAD(P)H, which occurs after hydrolysis and product dissociation. In conclusion, our work provides significant insight into ALDHs catalytic mechanism in bridging the gap between the well-studied chemical steps and a conformational transition essential for catalysis.…”

Section: ■ Discussionmentioning

confidence: 99%

“…By combining our past and present kinetic and structural studies with molecular dynamics simulations, ,, it is now possible to build up an integrated scenario of the sequence of events encompassed in deacylation (Figure ). As soon as hydride transfer occurs, the NMNH is allowed to flip through concerted up and down motion of the gatekeeper loop and reaches the hydrophobic exit pocket.…”

Section: Discussionmentioning

confidence: 99%

“…In ALDHs, local dynamics are associated with formation of an enzyme–substrate–cofactor ternary complex competent for acylation. This is the case, e.g., for nonphosphorylating glyceraldehyde 3-phosphate dehydrogenase (GAPN) that undergoes ligand-induced thermal destabilization and retinal dehydrogenase 2 (ALDH1A2) whose substrate specificity is ensured by a disorder to order transition. , In hydrolytic ALDHs, cofactor binding mode and motions control deacylation to some extent. − Indeed, the presence of multiple conformations of the nicotinamide mononucleotide (NMN) moiety is supported by the majority of the X-ray structures of ALDH-NAD(P) binary complexes − as well as NMR studies . During the acylation, the β-methyl group of the invariant conserved Thr244 residue allows the nicotinamide ring to adopt a productive conformation for hydride transfer (Figure B,C).…”

Section: Introductionmentioning

confidence: 99%

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Enzyme Active Site Loop Revealed as a Gatekeeper for Cofactor Flip by Targeted Molecular Dynamics Simulations and FRET-Based Kinetics

et al. 2019

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Structural motions are key events in enzyme catalysis, as exemplified by the conformational dynamics associated with the cofactor in the catalytic mechanism of hydrolytic NAD(P)-dependent aldehyde dehydrogenases. We previously showed that, after the oxidoreduction step, the reduced cofactor must adopt a flipped conformation, which positions the nicotinamide in a conserved cavity that might constitute the exit door for NAD(P)H. However, the molecular basis that make this movement possible is unknown. Based on the pre-and postflip Xray structures, targeted molecular dynamic simulations enabled us to identify the E 268 LGG 271 conserved loop that must shift to allow reduced nicotinamide conformational switch. To monitor cofactor movements within the active site, we used an intrinsic fluorescence resonance energy transfer signal between Trp177 and the reduced nicotinamide moiety to kinetically track the flip during the catalytic cycle of retinal dehydrogenase 2 (ALDH1A2). Decreasing loop flexibility by substituting Ala for Gly271 drastically reduced the rate constant associated with this movement that became rate-limiting. We thus propose that the E 268 LGG 271 loop acts as a gatekeeper for cofactor flipping. Similar approaches applied to a CoA-dependent aldehyde dehydrogenase showed that cofactor flipping likely extends to the whole ALDH family, thus bridging the gap between the well-studied chemical steps and a conformational transition essential for catalysis.

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Unraveling the function of paralogs of the aldehyde dehydrogenase super family from Sulfolobus solfataricus

et al. 2013

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Aldehyde dehydrogenases (ALDHs) have been well established in all three domains of life and were shown to play essential roles, e.g., in intermediary metabolism and detoxification. In the genome of Sulfolobus solfataricus, five paralogs of the aldehyde dehydrogenases superfamily were identified, however, so far only the non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase (GAPN) and α-ketoglutaric semialdehyde dehydrogenase (α-KGSADH) have been characterized. Detailed biochemical analyses of the remaining three ALDHs revealed the presence of two succinic semialdehyde dehydrogenase (SSADH) isoenzymes catalyzing the NAD(P)(+)-dependent oxidation of succinic semialdehyde. Whereas SSO1629 (SSADH-I) is specific for NAD(+), SSO1842 (SSADH-II) exhibits dual cosubstrate specificity (NAD(P)(+)). Physiological significant activity for both SSO-SSADHs was only detected with succinic semialdehyde and α-ketoglutarate semialdehyde. Bioinformatic reconstructions suggest a major function of both enzymes in γ-aminobutyrate, polyamine as well as nitrogen metabolism and they might additionally also function in pentose metabolism. Phylogenetic studies indicated a close relationship of SSO-SSALDHs to GAPNs and also a convergent evolution with the SSADHs from E. coli. Furthermore, for SSO1218, methylmalonate semialdehyde dehydrogenase (MSDH) activity was demonstrated. The enzyme catalyzes the NAD(+)- and CoA-dependent oxidation of methylmalonate semialdehyde, malonate semialdehyde as well as propionaldehyde (PA). For MSDH, a major function in the degradation of branched chain amino acids is proposed which is supported by the high sequence homology with characterized MSDHs from bacteria. This is the first report of MSDH as well as SSADH isoenzymes in Archaea.

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Adenine Binding Mode Is a Key Factor in Triggering the Early Release of NADH in Coenzyme A-dependent Methylmalonate Semialdehyde Dehydrogenase

Cited by 5 publications

References 39 publications

Retinoic acid biosynthesis catalyzed by retinal dehydrogenases relies on a rate-limiting conformational transition associated with substrate recognition

Retinoic acid biosynthesis catalyzed by retinal dehydrogenases relies on a rate-limiting conformational transition associated with substrate recognition

Enzyme Active Site Loop Revealed as a Gatekeeper for Cofactor Flip by Targeted Molecular Dynamics Simulations and FRET-Based Kinetics

Unraveling the function of paralogs of the aldehyde dehydrogenase super family from Sulfolobus solfataricus

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