The crystal structures of both isoforms of the aminoaldehyde dehydrogenase from pea (PsAMADH) have been solved recently [Tylichováet al. (2010) J Mol Biol396, 870-882]. The characterization of the PsAMADH2 proteins, altered here by site-directed mutagenesis, suggests that the D110 and D113 residues at the entrance to the substrate channel are required for high-affinity binding of ω-aminoaldehydes to PsAMADH2 and for enzyme activity, whereas N162, near catalytic C294, contributes mainly to the enzyme's catalytic rate. Inside the substrate cavity, W170 and Y163, and, to a certain extent, L166 and M167 probably preserve the optimal overall geometry of the substrate channel that allows for the appropriate orientation of the substrate. Unconserved W288 appears to affect the affinity of the enzyme for the substrate amino group through control of the substrate channel diameter without affecting the reaction rate. Therefore, W288 may be a key determinant of the differences in substrate specificity found among plant AMADH isoforms when they interact with naturally occurring substrates such as 3-aminopropionaldehyde and 4-aminobutyraldehyde.

• MeSH
• aldehydoxidoreduktasy chemie   genetika   izolace a purifikace   metabolismus   MeSH
• aldehydy metabolismus   MeSH
• aminokyseliny aromatické metabolismus   MeSH
• aminokyseliny dikarboxylové metabolismus   MeSH
• betain analogy a deriváty   metabolismus   MeSH
• biokatalýza MeSH
• cirkulární dichroismus MeSH
• hrách setý enzymologie   MeSH
• izoenzymy metabolismus   MeSH
• katalytická doména MeSH
• kinetika MeSH
• mutageneze cílená MeSH
• mutantní proteiny chemie   izolace a purifikace   metabolismus   MeSH
• propylaminy metabolismus   MeSH
• rekombinantní proteiny chemie   izolace a purifikace   metabolismus   MeSH
• rostlinné proteiny chemie   genetika   izolace a purifikace   metabolismus   MeSH
• sekundární struktura proteinů MeSH
• substrátová specifita MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Plant ALDH10 family members are aminoaldehyde dehydrogenases (AMADHs), which oxidize ω-aminoaldehydes to the corresponding acids. They have been linked to polyamine catabolism, osmoprotection, secondary metabolism (fragrance), and carnitine biosynthesis. Plants commonly contain two AMADH isoenzymes. We previously studied the substrate specificity of two AMADH isoforms from peas (PsAMADHs). Here, two isoenzymes from tomato (Solanum lycopersicum), SlAMADHs, and three AMADHs from maize (Zea mays), ZmAMADHs, were kinetically investigated to obtain further clues to the catalytic mechanism and the substrate specificity. We also solved the high resolution crystal structures of SlAMADH1 and ZmAMADH1a because these enzymes stand out from the others regarding their activity. From the structural and kinetic analysis, we can state that five residues at positions 163, 288, 289, 444, and 454 (PsAMADHs numbering) can, directly or not, significantly modulate AMADH substrate specificity. In the SlAMADH1 structure, a PEG aldehyde derived from the precipitant forms a thiohemiacetal intermediate, never observed so far. Its absence in the SlAMADH1-E260A structure suggests that Glu-260 can activate the catalytic cysteine as a nucleophile. We show that the five AMADHs studied here are capable of oxidizing 3-dimethylsulfoniopropionaldehyde to the cryo- and osmoprotectant 3-dimethylsulfoniopropionate. For the first time, we also show that 3-acetamidopropionaldehyde, the third aminoaldehyde besides 3-aminopropionaldehyde and 4-aminobutyraldehyde, is generally oxidized by AMADHs, meaning that these enzymes are unique in metabolizing and detoxifying aldehyde products of polyamine degradation to nontoxic amino acids. Finally, gene expression profiles in maize indicate that AMADHs might be important for controlling ω-aminoaldehyde levels during early stages of the seed development.

• MeSH
• aldehydoxidoreduktasy chemie   genetika   metabolismus   MeSH
• aldehydy chemie   MeSH
• chemické modely MeSH
• fylogeneze MeSH
• fyziologie rostlin MeSH
• kinetika MeSH
• krystalografie rentgenová metody   MeSH
• kukuřice setá enzymologie   MeSH
• mutageneze cílená MeSH
• NAD chemie   MeSH
• polyethylenglykoly chemie   MeSH
• regulace genové exprese enzymů * MeSH
• regulace genové exprese u rostlin * MeSH
• rostliny enzymologie   MeSH
• semena rostlinná metabolismus   MeSH
• Solanum lycopersicum enzymologie   MeSH
• substrátová specifita MeSH
• vazba proteinů MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH