Background: A non-catalytic vitamin B 6 biosynthesis protein (PDX1.2) exists in plants, but its role is unknown. Results: PDX1.2 interacts with its catalytic counterparts, enhancing their activity and forming a dodecameric complex induced under abiotic stress. Conclusion: PDX1.2 is a pesudoenzyme that regulates vitamin B 6 biosynthesis under abiotic stress. Significance: This work presents a model for regulation of vitamin B 6 biosynthesis and the existence/significance of pseudoenzymes in plants.
Ophidian l‐amino‐acid oxidase (l‐amino‐acid oxygen:oxidoreductase, deaminating, EC 1.4.3.2) is found in the venom of many poisonous snakes (crotalids, elapids and viperids). This FAD‐dependent glycoprotein has been studied from several snake species (e.g. Crotalus adamanteus, Crotalus atrox and Calloselasma rhodostoma) in detail with regard to the biochemical and enzymatic properties. The nature of glycosylation, however, as well as the chemical structure(s) of the attached oligosaccharide(s) are unknown. In view of the putative involvement of the glycan moiety in the biological effects of ophidian l‐amino‐acid oxidase, notably the apoptotic activity of the enzyme, structural knowledge is needed to evaluate its exact function. In this study we report on the glycosylation of l‐amino‐acid oxidase from the venom of the Malayan pit viper (Calloselasma rhodostoma). Its glycosylation is remarkably homogenous with the major oligosaccharide accounting for approximately 90% of the total sugar content. Based on detailed analysis of the isolated oligosaccharide by 2D NMR spectroscopies and MALDI‐TOF mass spectrometry the glycan is identified as a bis‐sialylated, biantennary, core‐fucosylated dodecasaccharide. The biological significance of this finding is discussed in light of the biological activities of the enzyme.
Thiamine (vitamin B 1 ) is ubiquitous and essential for cell energy supply in all organisms as a vital metabolic cofactor, known for over a century. In plants, it is established that biosynthesis de novo is taking place predominantly in green tissues and is furthermore limited to plastids. Therefore, transport mechanisms are required to mediate the movement of this polar metabolite from source to sink tissue to activate key enzymes in cellular energy generating pathways but are currently unknown. Similar to thiamine, polyamines are an essential set of charged molecules required for diverse aspects of growth and development, the homeostasis of which necessitates long-distance transport processes that have remained elusive. Here, a yeast-based screen allowed us to identify Arabidopsis (Arabidopsis thaliana) PUT3 as a thiamine transporter. A combination of biochemical, physiological, and genetic approaches permitted us to show that PUT3 mediates phloem transport of both thiamine and polyamines. Loss of function of PUT3 demonstrated that the tissue distribution of these metabolites is altered with growth and developmental consequences. The pivotal role of PUT3 mediated thiamine and polyamine homeostasis in plants, and its importance for plant fitness is revealed through these findings.
Thiamin pyrophosphate (TPP) is the active form of vitamin B 1 and works as an essential cofactor for enzymes in key metabolic pathways, such as the tricarboxylic acid (TCA) cycle and the pentose phosphate pathway. Although its action as a coenzyme has been well documented, the roles of TPP in plant metabolism are still not fully understood. Here, we investigated the functions of TPP in the regulation of the metabolic networks during photoperiod transition using previously described Arabidopsis (Arabidopsis thaliana) riboswitch mutant plants, which accumulate thiamin vitamers. The results show that photosynthetic and metabolic phenotypes of TPP riboswitch mutants are photoperiod dependent. Additionally, the mutants are more distinct from control plants when plants are transferred from a short-day to a long-day photoperiod, suggesting that TPP also plays a role in metabolic acclimation to the photoperiod. Control plants showed changes in the amplitude of diurnal oscillation in the levels of metabolites, including glycine, maltose, and fumarate, following the photoperiod transition. Interestingly, many of these changes are not present in TPP riboswitch mutant plants, demonstrating their lack of metabolic flexibility. Our results also indicate a close relationship between photorespiration and the TCA cycle, as TPP riboswitch mutants accumulate less photorespiratory intermediates. This study shows the potential role of vitamin B 1 in the diurnal regulation of central carbon metabolism in plants and the importance of maintaining appropriate cellular levels of thiamin vitamers for the plant's metabolic flexibility and ability to acclimate to an altered photoperiod.
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