In recent years, substantial progress has been made in exploring auxin conjugation and metabolism, primarily aiming at indole-3-acetic acid (IAA). However, the metabolic regulation of another key auxin, phenylacetic acid (PAA), remains largely uncharacterized. Here, we provide a comprehensive exploration of PAA metabolism in land plants. Through LC-MS screening across multiple plant species and their organs, we identified four previously unreported endogenous PAA metabolites: phenylacetyl-leucine, phenylacetyl-phenylalanine, phenylacetyl-valine, and phenylacetyl-glucose. Enzyme assays, genetic evidence, crystal structures, and docking studies demonstrate that PAA and IAA share core metabolic machinery, revealing a complex regulatory network that maintains auxin homeostasis. Furthermore, our study of PAA conjugation with amino acids and glucose suggests limited compensatory mechanisms within known conjugation pathways, pointing to the existence of alternative metabolic routes in land plants. These insights advance our knowledge of auxin-specific metabolic networks and highlight the unique complexity within plant hormone regulation.

• Klíčová slova
• Auxin, Gretchen Hagen 3, HPLC-MS/MS, conjugation, glucosyl ester, indole-3-acetic acid, metabolism, phenylacetic acid, plant,
• MeSH
• fenylacetáty * metabolismus   MeSH
• kyseliny indoloctové metabolismus   MeSH
• metabolické sítě a dráhy * MeSH
• regulátory růstu rostlin * metabolismus   MeSH
• vyšší rostliny * metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fenylacetáty * MeSH
• kyseliny indoloctové MeSH
• phenylacetic acid MeSH Prohlížeč
• regulátory růstu rostlin * MeSH

Adenosine undergoes ATP-dependent phosphorylation catalyzed by adenosine kinase (ADK). In plants, ADK also phosphorylates cytokinin ribosides, transport forms of the hormone. Here, we investigated the substrate preferences, oligomeric states, and structures of ADKs from moss (Physcomitrella patens) and maize (Zea mays) alongside metabolomic and phenotypic analyses. We showed that dexamethasone-inducible ZmADK overexpressor lines in Arabidopsis can benefit from a higher number of lateral roots and larger root areas under nitrogen starvation. We discovered that maize and moss enzymes can form dimers upon increasing protein concentration, setting them apart from the monomeric human and protozoal ADKs. Structural and kinetic analyses revealed a catalytically inactive unique dimer. Within the dimer, both active sites are mutually blocked. The activity of moss ADKs, exhibiting a higher propensity to dimerize, was 10-fold lower compared with maize ADKs. Two monomeric structures in a ternary complex highlight the characteristic transition from an open to a closed state upon substrate binding. This suggests that the oligomeric state switch can modulate the activity of moss ADKs and probably other plant ADKs. Moreover, dimer association represents a novel negative feedback mechanism, helping to maintain steady levels of adenosine and AMP.

• Klíčová slova
• Physcomitrella patens, Zea mays, Adenosine kinase, SnRK, crystal structure, cytokinin, overexpression, purine, riboside,
• MeSH
• adenosinkinasa * metabolismus   genetika   chemie   MeSH
• Arabidopsis genetika   enzymologie   metabolismus   MeSH
• kukuřice setá * enzymologie   genetika   metabolismus   MeSH
• mechy * enzymologie   genetika   metabolismus   MeSH
• multimerizace proteinu MeSH
• rostlinné proteiny * metabolismus   genetika   chemie   MeSH
• substrátová specifita MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• adenosinkinasa * MeSH
• rostlinné proteiny * MeSH

Aldehyde dehydrogenases (ALDHs) represent a superfamily of enzymes, which oxidize aldehydes to the corresponding acids. Certain families, namely ALDH9 and ALDH10, are best active with ω-aminoaldehydes arising from the metabolism of polyamines such as 3-aminopropionaldehyde and 4-aminobutyraldehyde. Plant ALDH10s show broad specificity and accept many different aldehydes (aliphatic, aromatic and heterocyclic) as substrates. This work involved the above-mentioned aminoaldehydes acylated with dicarboxylic acids, phenylalanine, and tyrosine. The resulting products were then examined with native ALDH10 from pea and recombinant ALDH7s from pea and maize. This investigation aimed to find a common efficient substrate for the two plant ALDH families. One of the best natural substrates of ALDH7s is aminoadipic semialdehyde carrying a carboxylic group opposite the aldehyde group. The substrate properties of the new compounds were demonstrated by mass spectrometry of the reaction mixtures, spectrophotometric assays and molecular docking. The N-carboxyacyl derivatives were good substrates of pea ALDH10 but were only weakly oxidized by the two plant ALDH7s. The N-phenylalanyl and N-tyrosyl derivatives of 3-aminopropionaldehyde were good substrates of pea and maize ALDH7. Particularly the former compound was converted very efficiently (based on the kcat/Km ratio), but it was only weakly oxidized by pea ALDH10. Although no compound exhibited the same level of substrate properties for both ALDH families, we show that these enzymes may possess more common substrates than expected.

• Klíčová slova
• Acylation, Aldehyde dehydrogenase, Aminoaldehyde, Docking, Enzyme, Substrate,
• MeSH
• aldehyddehydrogenasa * metabolismus   chemie   genetika   MeSH
• aldehydy * metabolismus   chemie   MeSH
• hrách setý * enzymologie   MeSH
• kinetika MeSH
• kukuřice setá * enzymologie   MeSH
• oxidace-redukce MeSH
• rostlinné proteiny metabolismus   chemie   genetika   MeSH
• simulace molekulového dockingu * MeSH
• substrátová specifita MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• aldehyddehydrogenasa * MeSH
• aldehydy * MeSH
• rostlinné proteiny MeSH

Heterokonts, Alveolata protists, green algae from Charophyta and Chlorophyta divisions, and all Embryophyta plants possess an aldehyde dehydrogenase (ALDH) gene named ALDH12. Here, we provide a biochemical characterization of two ALDH12 family members from the lower plant Physcomitrella patens and higher plant Zea mays. We show that ALDH12 encodes an NAD+-dependent glutamate γ-semialdehyde dehydrogenase (GSALDH), which irreversibly converts glutamate γ-semialdehyde (GSAL), a mitochondrial intermediate of the proline and arginine catabolism, to glutamate. Sedimentation equilibrium and small-angle X-ray scattering analyses reveal that in solution both plant GSALDHs exist as equilibrium between a domain-swapped dimer and the dimer-of-dimers tetramer. Plant GSALDHs share very low-sequence identity with bacterial, fungal, and animal GSALDHs (classified as ALDH4), which are the closest related ALDH superfamily members. Nevertheless, the crystal structure of ZmALDH12 at 2.2-Å resolution shows that nearly all key residues involved in the recognition of GSAL are identical to those in ALDH4, indicating a close functional relationship with ALDH4. Phylogenetic analysis suggests that the transition from ALDH4 to ALDH12 occurred during the evolution of the endosymbiotic plant ancestor, prior to the evolution of green algae and land plants. Finally, ALDH12 expression in maize and moss is downregulated in response to salt and drought stresses, possibly to maintain proline levels. Taken together, these results provide molecular insight into the biological roles of the plant ALDH12 family.

• Klíčová slova
• ALDH12, Physcomitrella patens, Zea mays, glutamate γ-semialdehyde, proline,
• MeSH
• aldehyddehydrogenasa chemie   MeSH
• fylogeneze MeSH
• krystalografie rentgenová metody   MeSH
• prolin chemie   MeSH
• rostliny chemie   MeSH
• substrátová specifita MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• aldehyddehydrogenasa MeSH
• prolin MeSH