Evidence for folate‐salvage reactions in plants

Abstract

SummaryFolates in vivo undergo oxidative cleavage, giving pterin and p-aminobenzoylglutamate (pABAGlu) moieties. These breakdown products are excreted in animals, but their fate is unclear in microorganisms and unknown in plants. As indirect evidence from this and previous studies strongly suggests that plants can have high folatebreakdown rates (approximately 10% per day), salvage of the cleavage products seems likely. Four sets of observations support this possibility. First, cleavage products do not normally accumulate: pools of pABAGlu (including its polyglutamyl forms) are equivalent to, at most, 4-14% of typical total folate pools in Arabidopsis thaliana, Lycopersicon esculentum and Pisum sativum tissues. Pools of the pterin oxidation end-product pterin-6-carboxylate are, likewise, fairly small (3-37%) relative to total folate pools. Second, little pABAGlu built up in A. thaliana plantlets when net folate breakdown was induced by blocking folate synthesis with sulfanilamide. Third, A. thaliana and L. esculentum tissues readily converted supplied breakdown products to folate synthesis precursors: pABAGlu was hydrolysed to p-aminobenzoate and glutamate, and dihydropterin-6-aldehyde was reduced to 6-hydroxymethyldihydropterin. Fourth, both these reactions were detected in vitro; the reduction used NADPH as cofactor. An alternative salvage route for pABAGlu, direct reincorporation into dihydrofolate via the action of dihydropteroate synthase, appears implausible from the properties of this enzyme. We conclude that plants are excellent organisms in which to explore the biochemistry of folate salvage.

Metabolism of the Folate Precursor p-Aminobenzoate in Plants

“…Because the mean yield of vacuoles was 10.1% as judged from vacuole numbers and ␣-mannosidase recovery (Fig. 5A), we can calculate that, on average, 79% (range 60 -90%) of the 14 C was located in vacuoles in vivo (Fig. 5B), this being a minimum estimate because 14 C may have leaked out during isolation.…”

“…5A), we can calculate that, on average, 79% (range 60 -90%) of the 14 C was located in vacuoles in vivo (Fig. 5B), this being a minimum estimate because 14 C may have leaked out during isolation. Of the 14 C label recovered from vacuoles, HPLC analysis demonstrated that essentially none was in the form of free pABA and that 80% was in the form of pABA-Glc (Fig.…”

“…Moreover, tests showed that omitting polyvinylpolypyrrolidone from our extraction buffer and cutting the 2-mercapto- 14 C]pABA dose given to leaflets used to prepare vacuoles. After the autoradiograph on the left was taken, the 14 C contents of TLC zones were determined, and expressed as percentages of the total 14 C label on the plate. The pABA-Glc zone (PG) contained 72% of the total label.…”

“…Pea (Pisum sativum cv. Laxton's Improved Progress 9) plants were grown as described (14). Cauliflower was obtained from a local market.…”

Nonflowering Plants Possess a Unique Folate-Dependent Phenylalanine Hydroxylase That Is Localized in Chloroplasts

“…The main plant pterins are the folate synthesis intermediates dihydroneopterin, dihydromonapterin, and hydroxymethyldihydropterin (Kohashi et al, 1980;Díaz de la Garza et al, 2004; Hossain et al, 2004;Orsomando et al, 2006). These might be reduced to the tetrahydro level by a dihydropterin reductase like those in bacteria or protists (Gourley et al, 2001;Pribat et al, 2010).…”