Phytoalexin Elicitor Activity Research Articles

ChemInform Abstract: Chemoselective Glycosylation Strategy for the Convergent Assembly of Phytoalexin-Elicitor Active Oligosaccharides and Their Photoreactive Derivatives.

A highly convergent route for the synthesis of branched glucosides having phytoalexin-elicitor activity has been developed. The readily available building blocks 2, 3, and 6 were used for the assembly of a core trisaccharide in two chemo- and regioselective glycosylations. Compound 7 was converted into the artificial spacer-containing glycosyl acceptor 9. Regioselective condensation of fragment 7 with 9 led to the formation of fully protected hexasaccharide 14. Deprotection of hexasaccharide 14 followed by the reaction with 4-azidosalicylate gave compound 1b. It was established that compound 1b is a photoreactive compound. Reaction time and conditions were established for photolysis of compound 1b and labeling with radioactive iodine. A soybean root binding site for an elicitor-active hepta-β-glucoside bound compound 1b with the same affinity as the underivatized hepta-β-glucoside.

Synthesis, (1→3)-β- d-glucanase-binding ability and phytoalexin-elicitor activity of ( R)-2,3-epoxypropyl (1→3)-β- d-pentaglucoside

The (1→3)-β- d-pentaglucoside was synthesized as its ( R)-2,3-epoxypropyl glycoside via 2 + 3 strategy. The disaccharide donor 8 was obtained by 3-selective coupling of 2 with 4, followed by deallylation, and trichloroacetimidation. Meanwhile, the trisaccharide acceptor 12 was prepared by coupling of 10 with 4, followed by deacetylation. Condensation of 8 with 12, followed by epoxidation, and deprotection, gave the target pentaoside. The results of these bioassays demonstrated that the (1→3)-β- d-glucanase was obviously inactivated by 15 with k app = 3.79 × 10 −4 min −1. At the same time, we found that the 15 was more active as compared to the laminaripentaose in eliciting phytoalexin accumulation in tobacco cotyledon tissue, and it could be kept longer time than laminaripentaose, which indicated it is much more stable than laminaripentaose.

Synthesis, (1 → 3)-β- D-glucanase-binding ability, and phytoalexin-elicitor activity of a mixture of 3,4-epoxybutyl (1 → 3)-β- D-oligoglucosides

We describe a approach for the synthesis of a mixture of 3,4-epoxybutyl (1 → 3)-β- D-oligoglucosides. The particular (1 → 3)-β- D-glucan isolated from the cell walls of Saccharomyces cerevisiae was recovered from the aqueous medium as water-insoluble particles by the spray drying (GS) method, and it was characterized by FTIR spectroscopy. The acid-solubilized (1 → 3)-β- D-oligoglucosides were prepared by partial acid hydrolysis of glucan particles, which were qualitatively analyzed by fluorophore-assisted carbohydrate electrophoresis (FACE). The peracetylated 3-butenyl (1 → 3)-β- D-oligoglucosides were synthesized by treating peracetylated (1 → 3)-β- D-oligoglucosides with the 3-butenyl alcohols and a Lewis acid (SnCl 4) catalyst. Epoxidation of the peracetylated 3-butenyl oligoglucosides took place with m-chloroperoxybenzoic acid ( m-CPBA). NaOMe in dry methanol was used for the deacetylation of the blocked derivatives, to give the 3,4-epoxybutyl (1 → 3)-β- D-oligoglucoside mixture in an overall yield of 21%. The sample was analyzed by positive-ion electrospray ionization mass spectrometry (ESIMS). In a 3,4-epoxybutyl (1 → 3)-β- D-oligoglucoside-binding (1 → 3)-β- D-glucanase assay, we found that the (1 → 3)-β- D-glucanase was obviously inactivated by the 3,4-epoxybutyl (1 → 3)-β- D-oligoglucosides. At the same time, we found the 3,4-epoxybutyl (1 → 3)-β- D-oligoglucoside mixture was more active as compared to the underivatized oligoglucoside mixture in eliciting phytoalexin accumulation in tobacco cotyledon tissue. Furthermore, it could be kept for a longer time than a (1 → 3)-β- D-oligoglucoside mixture, which indicated it is much more stable than (1 → 3)-β- D-oligoglucosides.

Convergent Total Syntheses of Oligosaccharides by One-Pot Sequential Stereoselective Glycosylations

Abstract A convergent total synthesis of F1α antigen, a member of the tumor-associated O-linked mucin glycosyl amino acid, was tried by one-pot sequential glycosylation. Highly α-selective glycosylation of amino acid 7 with thioglycoside 6 was successfully carried out by combining trityl trifluoromethanesulfonate (TrOTf) and N-iodosuccimide (NIS) which gave glycosyl amino acid 21 in high yield (97%, α/β = 83⁄17). Next, the glycosylation of thioglycoside 4 with galactosyl phenyl carbonate 2 or fluoride 3 was tried by the promotion of trityl tetrakis(pentafluorophenyl)borate [TrB(C6F5)4] or trifluoromethanesulfonic acid (TfOH); protected F1α 25 was afforded in 80 or 89% overall yield, respectively, by the further addition of glycosyl amino acid 5 and NIS. The desired trisaccharide was obtained in high yield after removal of the protecting groups. Next, a convergent total synthesis of branched hepta-β-glucoside 30 having phytoalexin-elicitor activity was efficiently performed by way of two one-pot sequential glycosylation reactions: that is, trisaccharide 34 was synthesized in high yield by TfOH-catalyzed one-pot glycosylation using three given monosaccharides (31, 35, and 36) as shown in Scheme 12 and by subsequent selective deprotection of 6′-O-TBDPS group. The second one-pot glycosylation of trisaccharide 34 with three monosaccharides (31, 32, and 33d) also proceeded smoothly to afford heptaglucoside 43 stereoselectively in 48% total yield based on monosaccharide 32. Phytoalexin-elicitor active branched hepta-β-glucoside 30 was afforded by the sequential deprotection.

Highly Efficient Syntheses of the Phytoalexin‐Elicitor Active β‐(1→3)‐Branched β‐(1→6)‐Linked Heptaglucose and Its Dodecyl Glycoside.

A highly efficient method for the synthesis of 3,6-branched gluco-oligosaccharides was developed by using 1,2:5,6-di-O-isopropylidene-α-D-glucofuranose, 2,3,4,6-tetra-O-henzoyl-α-D-glucopyranosyl trichloroacetimidate, and6-O-acetyl-2,3,4-tri-O-benzoyl-α-D-glucopyranosyl trichloroacetimidate through a regio- and stereoselective manner. The β-(1→3)-branched β-(1→6)-linked heptaglucose 1 and its dodecyl glycoside 2 having phytoalexin-elicitor activity were prepared using the developed strategy. Bioassays showed that the phytoalexin-elicitor activity of the dodecyl β-(1→3)-branched β-(1→6)-linked hexaglucoside 2 is slightly more than that of the corresponding reducing end free heptaglucose 1.

Highly Efficient Syntheses of the Phytoalexin-Elicitor Activeβ-(1→3)-Branched­β-(1→6)-Linked Heptaglucose and Its Dodecyl Glycoside

A highly efficient method for the synthesis of 3,6-branched gluco-oligosaccharides was developed by using 1,2:5,6-di-O-isopropylidene-α-d-glucofuranose, 2,3,4,6-tetra-O-benzoyl-α-d-glucopyranosyl trichloroacetimidate, and 6-O-acetyl-2,3,4-tri-O-benzoyl-α-d-glucopyranosyl trichloroacetimidate through a regio- and stereoselective manner. The β-(1→3)-branched β-(1→6)-linked heptaglucose 1 and its dodecyl glycoside 2 having phytoalexin-elicitor activity were prepared using the developed strategy. Bioassays showed that the phytoalexin-elicitor activity of the do­decyl β-(1→3)-branched β-(1→6)-linked hexaglucoside 2 is slightly more than that of the corresponding reducing end free heptaglucose 1.

Synthesis, protein-binding ability and phytoalexin-elicitor activity of epoxyalkyl (1-->3)-beta-D-oligoglucosides.

We describe a approach for the synthesis of (1-->3)-beta-D-oligosaccharide derivatives 10-18. 1-9 were synthesized by treating peracetylated (1-->3)-beta-D-oligosaccharides with the corresponding alkenyl alcohols and Lewis acid (SnCl(4)) catalyst. Epoxidation of the corresponding alkenyl oligoglucosides took place by m-CPBA. NaOMe in dry methanol was used for the deacetylation of the blocked derivatives, to give 10-18 in an overall yields of 25-32%. In subsequent glucan-binding protein of soybean assays, we found that 16 was most active, with an IC(50) value of 9 mM. However, the activities of 17, 18, 13, 14, 15, 10, 11, and 12 were gradually decreased. At the same time, we found 16 was most active as compared to the other (1-->3)-beta-D- oligoglucoside derivatives in eliciting phytoalexin accumulation in soybean cotyledon tissue, and 16 was kept longer time than (1-->3)-beta-D-glucohexaose, which indicated 16 is much more stable than (1-->3)-beta-D-glucohexaose.

Convergent and Convenient Total Synthesis of Phytoalexin‐Elicitor Active Heptasaccharide by One‐Pot Sequential Glycosylation.

Convergent and convenient total synthesis of branched hepta-β-glucoside 1 having phytoalexin-elicitor activity was efficiently accomplished by way of two one-pot sequential glycosylation reactions. Trisaccharide 8 was synthesized in high yield by TfOH-catalyzed one-pot glycosylation using three component monosaccharides and subsequent selective deprotection of a 6′-O-TBDPS group. The second one-pot glycosylation of trisaccharide 8 with the three monosaccharides smoothly proceeded to afford heptaglucoside 11 stereoselectively in 48% total yield based on monosaccharide 3. The targeted compound 1 was obtained in high yield after the removal of the protecting groups.

Synthesis of di-branched heptasaccharide by one-pot glycosylation using seven independent building blocks.

[reaction: see text] We describe an efficient synthesis of di-branched heptasaccharide 1 having phytoalexin elicitor activity in soybeans by one-pot glycosylation. The synthesis involves chemo- and regioselective sequential six-step glycosylations using seven independent building blocks and sequential removal of acyl- and benzyl ether-type protecting groups. The coupling of seven building blocks requires only four chemoselective activitable leaving groups of glycosyl donors. Both the glycosylation and deprotection reactions can be achieved utilizing a parallel manual synthesizer.

Convergent and Convenient Total Synthesis of Phytoalexin-Elicitor Active Heptasaccharide by One-Pot Sequential Glycosylation

Abstract Convergent and convenient total synthesis of branched hepta-β-glucoside 1 having phytoalexin-elicitor activity was efficiently accomplished by way of two one-pot sequential glycosylation reactions. Trisaccharide 8 was synthesized in high yield by TfOH-catalyzed one-pot glycosylation using three component monosaccharides and subsequent selective deprotection of a 6′-O-TBDPS group. The second one-pot glycosylation of trisaccharide 8 with the three monosaccharides smoothly proceeded to afford heptaglucoside 11 stereoselectively in 48% total yield based on monosaccharide 3. The targeted compound 1 was obtained in high yield after the removal of the protecting groups.

Solid-Phase Synthesis of a Phytoalexin Elicitor-Active Tetraglucosyl Glucitol

A solid-phase synthesis of pentasaccharide 1 that has phytoalexin elicitor activity for rice is described. Efficient block-type synthesis by β-selective glycosylation with thioglycosides A 3 and B 2 at the 3-position of the polymer-supported glycosyl acceptor has been demonstrated.

The first synthesis of tetraglucosyl glucitol having phytoalexin-elicitor activity in rice cells based on a sequential glycosylation strategy

We describe a highly convergent approach for the synthesis of the tetraglucosyl glucitol 1 and its derivatives 4 , 5 and 6 which exhibit phytoalexin-elicitor activity in rice cells based on sequential glycosylation.

Chemoselective Glycosylation Strategy for the Convergent Assembly of Phytoalexin-Elicitor Active Oligosaccharides and Their Photoreactive Derivatives

A highly convergent route for the synthesis of branched glucosides having phytoalexin-elicitor activity has been developed. The readily available building blocks 2, 3, and 6 were used for the assembly of a core trisaccharide in two chemo- and regioselective glycosylations. Compound 7 was converted into the artificial spacer-containing glycosyl acceptor 9. Regioselective condensation of fragment 7 with 9 led to the formation of fully protected hexasaccharide 14. Deprotection of hexasaccharide 14 followed by the reaction with 4-azidosalicylate gave compound 1b. It was established that compound 1b is a photoreactive compound. Reaction time and conditions were established for photolysis of compound 1b and labeling with radioactive iodine. A soybean root binding site for an elicitor-active hepta-β-glucoside bound compound 1b with the same affinity as the underivatized hepta-β-glucoside.

Synthesis of glycopeptides with phytoalexin elicitor activity — III. Syntheses of hexaglycosyl hexapeptides and a nonaglycosyl hexapeptide

A block synthesis of the model compound for the phytoalexin elicitor-active glycoprotein is described. Combination of the C-terminus free compounds, N-(9- fluorenylmethoxycarbonyl)-O-(tert- butyl)- l- seryl- l- proline (1) or N-(9- fluorenylmethoxycar bonyl)-(2,3,4,6- tetra-O- acetyl-β- d- glucopyranosyl)-(1→ 6)-(2,3,4- tri-O- acetyl-α- d- mannopyranosyl)-(1→ 6)-(2,3,4- tri-O- acetyl-α- d- mannopyranosyl)- l- seryl- l- proline (2) with the N-terminus free compounds, 2,3,4,6- tetra-O- acetyl-β- d- glucopyranosyl(1→6)-(2,3,4- tri-O- acetyl-α- d- mannopyranosyl)-(1→6)-(2,3,4- tri-O- acetyl-α- d- mannopyranosyl)- l- seryl- l- prolyl- l- seryl- l- proline methyl ester (4), O-(tert- butyl)- l- seryl- l- prolyl-(2,3,4,6- tetra-O- acetyl- β- d- glucopyranosyl)- (1→6)-(2,3,4- tri-O- acetyl-α- d- mannopyranosyl)-(1→6)-(2,3,4- tri-O- acetyl-α- d- mannopyranosyl)- l- seryl- l- proline methyl ester (6) or 2,3,4,6- tetra-O- acetyl-β- d- glucopyranosyl-(1→6)-(2,3,4- tri-O- acetyl-α- d- mannopyranosyl)-(1→6)-(2,3,4- tri-O- acetyl-α- d- mannopyranosyl)- l- seryl- l- prolyl-(2,3,4,6- tetra-O- acetyl-β- d- glucopyranosyl)-(1→6)-(2,3,4- tri-O- acetyl-α- d- mannopyranosyl)-(1→6)-(2,3,4- tri- O- acetyl-α- d- mannopyranosyl)- l- seryl- l- proline methyl ester (8), by use of N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) gave three hexaglycosyl hexapeptides and a nonaglycosyl hexapeptide derivatives (9, 11, 14, and 17). These N-terminus free compounds were derived from triglycosyl tetrapeptides (3, and 5) or a hexaglycosyl tetrapeptide (7) on selective deblock reaction by morpholine. The hexaglycosyl hexapeptides (10, 13, and 16) and the nonaglycosyl hexapeptide (18) have been prepared by the convergent block synthesis.

Synthesis of a Methyl Heptaglucoside with Phytoalexin Elicitor Activity Based on Oxidative Coupling of Glucals

AbstractSeveral suitable building blocks for the construction of the phytoalexin elicitor α‐methyl‐32, 34‐di‐β‐D‐glucopyranosylgentiopentaoside (2) were readily accessible by oxidative coupling of glucals. Block coupling of trimeric phenylseleno‐ and ethylthioglucosyl donors 17 and 18 with tetrasaccharide 16 in the presence of the thiophilic promoter N‐iodosuccinimide and catalytic trifluoromethanesulfonic acid furnished the desired heptaglucan 2 in high overall yield.

Induction of resistance mechanisms in barley by yeast‐derived elicitors

SummaryYeast cell‐wall extracts (YE) had high phytoalexin elicitor activity on soybeans and induced resistance against barley powdery mildew in the normally highly susceptible cv. Golden Promise. Following treatment with YE there was rapid stimulation of phenylalanine ammonia‐lyase (PAL) activity and faster formation of papillae in response to attempted fungal penetration. The spray adjuvants Agral and LI‐700 each had poor phytoalexin elicitor activity and caused a slight enhancement of papilla formation and a low level of mildew control. The addition of the adjuvant to YE did not affect phytoalexin elicitation on soybeans but significantly enhanced the speed and the extent of PAL activity, papilla formation and mildew control. It is proposed that this is due to greater coverage and improved persistence enabling increased uptake of YE into the barley leaves.

Polymer‐supported solution synthesis of a heptaglucoside having phytoalexin elicitor activity

AbstractAn efficient synthesis of methyl heptaglucoside 2 using poly(ethyleneglycol)monomethyl ether as a polymer support is described. Elongation of the acceptor 11, anchored via a succinoyl linkage to PEG, with the appropriate glycosyl donors 4–6 and intermittent protective group manipulations gave fully protected heptamer 18. Deprotection of 18 furnished, after purification as its peracetate, title compound 2 in a good overall yield.

Characterization of two Sclerotinia sclerotiorum polygalacturonases with different abilities to elicit glyceollin in soybean

Two endo-polygalacturonase isoenzymes (PG-II and PG-IV), with masses of 34 and 30 kDa respectively, were purified from soybean hypocotyls infected by Sclerotinia sclerotiorum. The pH optimum for both isoenzymes was about 4.6, but PG-IV exhibited a broader range of pH activity. PG-IV showed a much higher affinity for pectin than did PG-II. PG-II hydrolyzed polygalacturonic acid in a more random fashion than PG-IV. Oligouronides produced by PG-II showed a higher phytoalexin elicitor activity. PG-IV produced a large degree of maceration of soybean hypocotyls releasing a significant amount of uronides. The properties of PG-II and PG-IV are discussed in relation to the different ability of the two isoenzymes to elicit glyceollin in soybean.

Bacterial endopectate lyase: evidence that plant cell wall pH prevents tissue maceration and increases the half-life of elicitor-active oligogalacturonides

The abilities of endopectate lyase (α-1,4-poly- d-galacturonide lyase), purified from the bacterial phytopathogen Erwinia carotovora, to macerate potato tissue and to produce elicitor-active oligogalacturonides from polygalacturonic acid were studied at various pH values. The rate at which the lyase depolymerized polygalacturonic acid was maximum (100%) at pH 9·0, 10% at pH 8·0, and 3% at pH 7·0. Endopectate lyase macerated potato medullary tissue discs extensively at pH 9·0, to a lesser extent between pH 8·0 and 7·0, and did not detectably macerate the discs at pH values lower than 7·0 or in water (pH approx. 6·0). Digestion products of polygalacturonic acid treated with endopectate lyase at pH 5·75, the experimentally determined pH of soybean cotyledon exudates, and at pH 8·5, optimum for polygalacturonic acid depolymerization, were tested for phytoalexin elicitor activity in the soybean cotyledon bioassay. The elicitor activity of the digestion products obtained at pH 5·75 increased during the first 40 h of incubation and slowly decreased thereafter as degradation to short elicitor-inactive oligomers progressed. On the other hand, the activity of the digestion products obtained at pH 8·5 dropped to a very low level in less than 20 min. The stability of the oligogalacturonides with degrees of polymerization between 10 and 20, i.e. the oligogalacturonides which exhibit high elicitor activity, was longer than 24 h when the digestion of polygalacturonic acid was carried out at pH 5·75, but was less than 20 min when digestions were carried out at pH 8·5. These studies suggest that, in vivo, endopectate lyases are unlikely to hydrolyse plant cell wall pectin extensively or cause maceration at early stages of pathogenesis, but that the lyases are able to generate elicitor-active oligogalacturonides.

Substrate Degradation Patterns of Polygalacturonic Acid Lyase fromErwinia carotovoraandBacillus polymyxaand Release of Phytoalexin-eliciting Oligosaccharides from Potato Cell Walls

The patterns of substrate degradation by purified pectate lyase (PGL) (E.C. 4.2.2.2) from Erwinia carotovora and Bacillus polymyxa were compared. Reaction products released by both enzymes from potato cell walls, sodium polypectate and citrus pectin were separated by anion exchange chromatography using a TSK DEAE-5PW column and measured quantitatively. The relative amounts of oligomers released by both enzymes varied, especially the level of unsaturated tetramers. Degradation patterns also varied according to the substrate used and results with citrus pectin suggested that methylation reduced the ability of E. carotovora PGL to release wall fragments. Oligomers released from potato cell walls by E. carotovora PGL were pooled separately and assayed for phytoalexin elicitor activity using the soybean cotyledon bioassay. Fractions containing deca- and undecagalacturonides had the highest elicitor activity when tested at 5.0μg of uronic acid per cotyledon.