White Lupin Roots Research Articles

Farnesyl Pyrophosphate Synthase from White Lupin: Molecular Cloning, Expression, and Purification of the Expressed Protein

Plants produce a variety of sesquiterpenoid compounds with diverse biological functions, whose synthesis is initiated by farnesyl pyrophosphate synthase [EC 2.5.1.1, EC 2.5.1.10], The lack of availability of molecular tools to analyze this enzyme has, thus far, prevented the study of its expression and regulation in plants, A DNA fragment corresponding to a portion of the farnesyl pyrophosphate synthase gene was amplified by the polymerase chain reaction using degenerate primers designed from two highly conserved domains (FLV(A/L)DD(I/M)MD and FQIQDD-DYLD) found in eukaryotic farnesyl pyrophosphate synthase sequences. A clone, pS19, of a 438-bp PCR fragment was used to screen a white lupin root cDNA library, Two full-length cDNA clones ( pFPS1 and pFPS2) were isolated and sequenced, and one of them ( pFPS2) was expressed in a bacterial system and the enzyme protein encoded by the clone was purified, The 1345-bp insert of pFPS2 contains a 1026-bp open reading frame, encoding a 342-amino-acid peptide with a calculated molecular mass of 39,310 Da. The deduced amino acid sequence of lupin farnesyl pyrophosphate synthase pFPS2 shares 90 and 79% identity with those from Lupinus albus ( pFPSI) and Arabidopsis thaliana, respectively, 51% with the yeast enzyme, and 44% identity with those from rat and human. The overexpressed protein, which was purified to near homogeneity, displayed both dimethylalkyl transferase and geranyl transferase activities. Polyclonal antibodies raised against the purified protein immunorecognized a ca 39-kDa protein in lupin root extracts.

Role of Nitrogen and Plant Growth Regulators in the Exudation and Accumulation of lsoflavonoids by Roots of Intact White (Lupin ( Lupinus albus L.) Plants

Summary Two isoflavonoid glycosides (7-O-β-glucopyranosides of 2′-hydroxygenistein and genistein) and up to six aglycones (2′-hydroxygenistein, genistein, wighteone, luteone, lupinisoflavone A, parvisoflavone B) were identified in root exudates and/or root extracts from white lupin ( Lupinus albus L. cv. Bac) plants. Three-d-old seedlings were cultivated for 14d on 1/2 Knop's nutrient solutions. Various treatments, differing in the form (No 3 − , NH 4 + , NH 4 N0 3 , urea) and concentration of nitrogen, and the presence of plant growth regulators in the medium were tested. Nitrogen, in all forms applied, affects both the exudation and accumulation of phenolics with the inhibitory activites arranged in the order: N0 3 − > NH 4 N0 3 > urea > NH 4 + . With higher concentration of N in the medium, the exudation of phenolics decreases. Processes of isoflavone exudation and accumulation respond differentially to nitrogen status. Exogenously applied plant growth regulators modify inhibitory effects of nitrate on the exudation and accumulation of isoflavones. Their activity depend on the type of growth regulator used (NAA, BAP, GA 3 ), its chemical structure (IAA, IBA, NAA) and concentration in the medium. Lupin isoflavones might be potential signals in legume — Rhizobium interactions. These data show a correlation between plant nutrient status and the accumulation and exudation of putative phenolic signal molecules, thus indicating the existence of possible additional controls in the regulation of nodulation process.

Further Isoflavonoids from White Lupin Roots

Abstract Further investigation of the isoflavonoids in white lupin roots has revealed a new coumaronochromone with a 2-hydroxy-3-methyl-3-butenyl side chain (lupinalbin G), three new pyrano-isoflavones (lupinisoflavones K and L, and allolicoisoflavone B) and two new di-hydrofurano-isoflavones each with a 2,3-dihydroxy-3-methylbutyl substituent (lupinisoflavones M and N). The biogenetic pathways for these and other lupin isoflavones are briefly discussed.

Citric acid excretion and precipitation of calcium citrate in the rhizosphere of white lupin (Lupinus albus L.)

Abstract. White lupin (Lupinus albus L.) was grown for 13 weeks in a phosphorus (P) deficient calcareous soil (20% CaCO3, pH(H2O)7.5) which had been sterilized prior to planting and fertilized with nitrate as source of nitrogen. In response to P deficiency, proteoid roots developed which accounted for about 50% of the root dry weight. In the rhizosphere soil of the proteoid root zones, the pH dropped to 4.8 and abundant white precipitates became visible. X‐ray spectroscopy and chemical analysis showed that these precipitates consisted of calcium citrate. The amount of citrate released as root exudate by 13‐week‐old plants was about 1 g plant−1, representing about 23% of the total plant dry weight at harvest. In the rhizosphere soil of the proteoid root zones the concentrations of available P decreased and of available Fe, Mn and Zn increased. The strong acidification of the rhizosphere and the cation/anion uptake ratio of the plants strongly suggests that proteoid roots of white lupin excrete citric acid, rather than citrate, into the rhizosphere leading to intensive chemical extraction of a limited soil volume. In a calcareous soil, citric acid excretion leads to dissolution of CaCO3 and precipitation of calcium citrate in the zone of proteoid roots.

Stereochemical studies on dihydrofurano-isoflavones.

The stereochemistry of some dihydrofurano-isoflavones previously isolated from white lupin roots, or obtained following fungal metabolism of prenylated isoflavones, was investigated using CD spectroscopy. The osmate ester/pyridine complex of dextrorotatory lupinisoflavone A (1) exhibited a positive CD Cotton effect at 480nm, indicating a side-structure configuration (S at C-2''), opposite to that of natural rotenone (9), which afforded a negative Cotton effect at 474nm (R-configuration at C-2' on the side structure [ring E]). The stereochemistry of the laevorotatory luteone metabolite BC-1 (2) and lupinisoflavone D (4) (both R-configuration at C-2'') was similarly determined after converting to the corresponding dehydrate (10) or trimethyl-dehydrate (1b, 10a).

Stereochemical Studies on Dihydrofurano-isoflavones

The stereochemistry of some dihydrofurano-isoflavones previously isolated from white lupin roots, or obtained following fungal metabolism of prenylated isoflavones, was investigated using CD spectroscopy. The osmate ester/pyridine complex of dextrorotatory lupinisoflavone A (1) exhibited a positive CD Cotton effect at 480 nm, indicating a side-structure configuration (S at C- 2″), opposite to that of natural rotenone (9), which afforded a negative Cotton effect at 474 nm (R- configuration at C-2′ on the side structure [ring E]). The stereochemistry of the laevorotatory luteone metabolite BC-1 (2) and lupinisoflavone D (4) (both ^-configuration at C-2″) was similarly determined after converting to the corresponding dehydrate (10) or trimethyl-dehydrate (1b, 10a).

New coumaronochromones from white lupin, Lupinus albus L. (Leguminosae).

A further investigation of the isoflavonoid constituents occurring in roots of the white lupin (Lupinus albus L. cv. Kievskij Mutant) has yielded five new Coumaronochromones named lupinalbin A (1a), B (2a), C (3), D (4) and E (5). These isoflavonoids were identified by physicochemical methods involving the use of biogenetically related 2'-hydroxyisoflavones as reference compounds. The presence of the rare dihydrofurano-isoflavone, erythrinin C (16), in white lupin roots has also been established.

Fungitoxic dihydrofuranoisoflavones and related compounds in white lupin, Lupinus albus

Chromatographic investigation of a methanolic extract of white lupin roots has revealed the presence of six new dihydrofuranoisoflavones (lupinisoflavones A-F). Three monoprenylated (3,3-dimethylallyl-substituted) isoflavones (wighteone, luteone and licoisoflavone A), two diprenylated isoflavones [6,3′-di(3,3-dimethylallyl)genistein (lupalbigenin) and 6,3′-di(3,3-dimethylallyl)-2′-hydroxygenistein (2′-hydroxylupalbigenin)] and two pyranoisoflavones (parvisoflavone B and licoisoflavone B) have also been isolated from the same source. In addition to genistein, leaf extracts of L. italbus contain 3′- O-methylorobol which is presumed to be the precursor of lupisoflavone [5,7,4′-trihydroxy-3′-methoxy-6-(3,3-dimethylallyl)isoflavone]. Probable biogenetic relationships between the prenylated, and dihydrofurano-and pyrano-substituted isoflavones in roots and leaves of L. albus are briefly discussed.