Cell Walls Of Vascular Plants Research Articles

Lignin modification improves the biofuel production potential in transgenic Populus tomentosa

Lignin has been recognized for its negative impact on forage digestibility, tree pulping properties, and cellulosic biofuel production, although it is the major structural component of the secondarily thickened cell walls of vascular plants. Earlier studies have demonstrated that lignin modification improves forage digestibility and poplar pulping properties. To determine whether lignin modification has beneficial effect on saccharification of lignocellulosic biomass, we pretreated and then enzymatically hydrolyzed the mature wood from transgenic poplar plants that expressed the antisense transgenes of monolignol biosynthesis genes 4-coumarate: CoA ligase (4CL) or caffeoyl CoA 3-O-methyltransferase (CCoAOMT). Firstly, a long-term field trial was set up for the transgenic plants. Over five years, the reduced trend of lignin content remained stable in all transgenic lines. And a total lignin reduction of up to 10% did not alter the growth rate or biomass yield of the transgenic poplars. In the mature wood, suppression of CCoAOMT increased saccharification potential, but 4CL down-regulation had no significantly positive effect on saccharification. Sugar yield were negatively correlated with soluble lignin content of dried, extractive-free stem biomass. These results imply that lignin modification can facilitate the process of saccharification for biofuel production in tree crops.

The ability of land plants to synthesize glucuronoxylans predates the evolution of tracheophytes

Glucuronoxylans with a backbone of 1,4-linked β-D-xylosyl residues are ubiquitous in the secondary walls of gymnosperms and angiosperms. Xylans have been reported to be present in hornwort cell walls, but their structures have not been determined. In contrast, the presence of xylans in the cell walls of mosses and liverworts remains a subject of debate. Here we present data that unequivocally establishes that the cell walls of leafy tissue and axillary hair cells of the moss Physcomitrella patens contain a glucuronoxylan that is structurally similar to glucuronoxylans in the secondary cell walls of vascular plants. Some of the 1,4-linked β-D-xylopyranosyl residues in the backbone of this glucuronoxylan bear an α-D-glucosyluronic acid (GlcpA) sidechain at O-2. In contrast, the lycopodiophyte Selaginella kraussiana synthesizes a glucuronoxylan substituted with 4-O-Me-α-D-GlcpA sidechains, as do many hardwood species. The monilophyte Equisetum hyemale produces a glucuronoxylan with both 4-O-Me-α-D-GlcpA and α-D-GlcpA sidechains, as does Arabidopsis. The seedless plant glucuronoxylans contain no discernible amounts of the reducing-end sequence that is characteristic of gymnosperm and eudicot xylans. Phylogenetic studies showed that the P. patens genome contains genes with high sequence similarity to Arabidopsis CAZy family GT8, GT43 and GT47 glycosyltransferases that are likely involved in xylan synthesis. We conclude that mosses synthesize glucuronoxylan that is structurally similar to the glucuronoxylans present in the secondary cell walls of lycopodiophytes, monilophytes, and many seed-bearing plants, and that several of the glycosyltransferases required for glucuronoxylan synthesis evolved before the evolution of tracheophytes.

Microbial degradation of lignin: how a bulky recalcitrant polymer is efficiently recycled in nature and how we can take advantage of this

SummaryLignin is the second most abundant constituent of the cell wall of vascular plants, where it protects cellulose towards hydrolytic attack by saprophytic and pathogenic microbes. Its removal represents a key step for carbon recycling in land ecosystems, as well as a central issue for industrial utilization of plant biomass. The lignin polymer is highly recalcitrant towards chemical and biological degradation due to its molecular architecture, where different non‐phenolic phenylpropanoid units form a complex three‐dimensional network linked by a variety of ether and carbon–carbon bonds. Ligninolytic microbes have developed a unique strategy to handle lignin degradation based on unspecific one‐electron oxidation of the benzenic rings in the different lignin substructures by extracellular haemperoxidases acting synergistically with peroxide‐generating oxidases. These peroxidases posses two outstanding characteristics: (i) they have unusually high redox potential due to haem pocket architecture that enables oxidation of non‐phenolic aromatic rings, and (ii) they are able to generate a protein oxidizer by electron transfer to the haem cofactor forming a catalytic tryptophanyl‐free radical at the protein surface, where it can interact with the bulky lignin polymer. The structure–function information currently available is being used to build tailor‐made peroxidases and other oxidoreductases as industrial biocatalysts.

Soft X-ray induced chemical modification of polysaccharides in vascular plant cell walls

Scanning transmission X-ray microscopy and micro carbon X-ray Absorption Near Edge Spectroscopy (C-XANES) can provide quantitative information regarding the distribution of the biopolymers cellulose, hemicellulose, and lignin in vascular plant cell walls. In the case of angiosperms, flowering plants, C-XANES may also be able to distinguish variations in lignin monomer distributions throughout the cell wall. Polysaccharides are susceptible to soft X-ray irradiation induced chemical transformations that may complicate spectral analysis. The stability of a model polysaccharide, cellulose acetate, to variable doses of soft X-rays under conditions optimized for high quality C-XANES spectroscopy was investigated. The primary chemical effect of soft X-ray irradiation on cellulose acetate involves mass loss coincident with de-acetylation. A lesser amount of vinyl ketone formation also occurs. Reduction in irradiation dose via defocusing does enable high quality pristine spectra to be obtained. Radiation induced chemical modification studies of oak cell wall reveals that cellulose and hemicellulose are less labile to chemical modification than cellulose acetate. Strategies for obtaining pristine C-XANES spectra of polysaccharides are presented.

Improvement of biomass through lignin modification

Lignin, a major component of the cell wall of vascular plants, has long been recognized for its negative impact on forage quality, paper manufacturing, and, more recently, cellulosic biofuel production. Over the last two decades, genetic and biochemical analyses of brown midrib mutants of maize, sorghum and related grasses have advanced our understanding of the relationship between lignification and forage digestibility. This work has also inspired genetic engineering efforts aimed at generating crops with altered lignin, with the expectation that these strategies would enhance forage digestibility and/or pulping efficiency. The knowledge gained from these bioengineering efforts has greatly improved our understanding of the optimal lignin characteristics required for various applications of lignocellulosic materials while also contributing to our understanding of the lignin biosynthetic pathway. The recent upswing of interest in cellulosic biofuel production has become the new focus of lignin engineering. Populus trichocarpa and Brachypodium distachyon are emerging as model systems for energy crops. Lignin research on these systems, as well as on a variety of proposed energy crop species, is expected to shed new light on lignin biosynthesis and its regulation in energy crops, and lead to rational genetic engineering approaches to modify lignin for improved biofuel production.

A Barley Xyloglucan Xyloglucosyl Transferase Covalently Links Xyloglucan, Cellulosic Substrates, and (1,3;1,4)-β-D-Glucans

Molecular interactions between wall polysaccharides, which include cellulose and a range of noncellulosic polysaccharides such as xyloglucans and (1,3;1,4)-beta-D-glucans, are fundamental to cell wall properties. These interactions have been assumed to be noncovalent in nature in most cases. Here we show that a highly purified barley xyloglucan xyloglucosyl transferase HvXET5 (EC 2.4.1.207), a member of the GH16 group of glycoside hydrolases, catalyzes the in vitro formation of covalent linkages between xyloglucans and cellulosic substrates and between xyloglucans and (1,3;1,4)-beta-D-glucans. The rate of covalent bond formation catalyzed by HvXET5 with hydroxyethylcellulose (HEC) is comparable with that on tamarind xyloglucan, whereas that with (1,3; 1,4)-beta-D-glucan is significant but slower. Matrix-assisted laser desorption ionization time-of-flight mass spectrometric analyses showed that oligosaccharides released from the fluorescent HEC:xyloglucan conjugate by a specific (1,4)-beta-D-glucan endohydrolase consisted of xyloglucan substrate with one, two, or three glucosyl residues attached. Ancillary peaks contained hydroxyethyl substituents (m/z 45) and confirmed that the parent material consisted of HEC covalently linked with xyloglucan. Similarly, partial hydrolysis of the (1,3;1,4)-beta-D-glucan:xyloglucan conjugate by a specific (1,3;1,4)-beta-D-glucan endohydrolase revealed the presence of a series of fluorescent oligosaccharides that consisted of the fluorescent xyloglucan acceptor substrate linked covalently with 2-6 glucosyl residues. These findings raise the possibility that xyloglucan endo-transglucosylases could link different polysaccharides in vivo and hence influence cell wall strength, flexibility, and porosity.

Raman imaging to investigate ultrastructure and composition of plant cell walls: distribution of lignin and cellulose in black spruce wood (Picea mariana)

A detailed understanding of the structural organization of the cell wall of vascular plants is important from both the perspectives of plant biology and chemistry and of commercial utilization. A state-of-the-art 633-nm laser-based confocal Raman microscope was used to determine the distribution of cell wall components in the cross section of black spruce wood in situ. Chemical information from morphologically distinct cell wall regions was obtained and Raman images of lignin and cellulose spatial distribution were generated. While cell corner (CC) lignin concentration was the highest on average, lignin concentration in compound middle lamella (CmL) was not significantly different from that in secondary wall (S2 and S2-S3). Images generated using the 1,650 cm(-1) band showed that coniferaldehyde and coniferyl alcohol distribution followed that of lignin and no particular cell wall layer/region was therefore enriched in the ethylenic residue. In contrast, cellulose distribution showed the opposite pattern-low concentration in CC and CmL and high in S2 regions. Nevertheless, cellulose concentration varied significantly in some areas, and concentrations of both lignin and cellulose were high in other areas. Though intensity maps of lignin and cellulose distributions are currently interpreted solely in terms of concentration differences, the effect of orientation needs to be carefully considered to reveal the organization of the wood cell wall.

Lignin Radicals in the Plant Cell Wall Probed by Kerr-Gated Resonance Raman Spectroscopy

Lignin radicals are crucial intermediates for lignin biosynthesis in the cell wall of vascular plants. In this work they were for the first time, to our knowledge, selectively observed in wood cell walls by laser-based Kerr-gated resonance Raman spectroscopy, and the observations were supported by density functional theory prediction of their vibrational properties. For dry wood cells a lignin radical Raman band is observed at 1570 cm −1 irrespective of species. For wet beech cells they were generated in situ and observed at 1606 cm −1. DFT/B3LYP/6-31+G(d) modeling results support that in beech they are formed from syringyl (S) phenolic moieties and in spruce from guaiacyl (G) phenolic moieties. The observed lignin radical band is predicted as G is ∼1597 cm −1 and S is ∼1599 cm −1, respectively, and is assigned the (Wilson notation) ν 8a phenyl ring mode. The RR band probes lignin radical properties, e.g., spin density distribution, and these respond to charge polarization or hydrogen bonding to proximate water molecules. These observations can be crucial for an understanding of the factors that control cell wall structure during biosynthesis of vascular plants and demonstrate the unique potential of RR spectroscopy of lignin radicals.

Dehydrogenative polymerization of conifery alcohol on macromolecular lignin templates

Lignins, the aromatic biopolymers present in all vascular plant cell walls, may embody as many as 10 different linkages between the constituent ( p-hydroxyphenyl)propane residues. The final step in lignin biosynthesis has traditionally been thought to involve the random coupling of radical intermediates produced during the dehydrogenative polymerization of monolignol precursors. However, the enzyme-catalysed dehydropolymerization of coniferyl alcohol in homogeneous solution has now been found to be strongly affected by small quantities of macromolecular lignin components. Under these circumstances, when the radical concentrations remained low, the molecular weight distributions of the dehydropolymerisates formed with horseradish peroxidase and H 2O 2 exhibited dramatic increases in the populations of the largest species. The phenomenon was not suppressed by prior methylation of the aromatic hydroxyl groups on the causative lignin macromolecules—which, therefore, were able to promote the formation of large covalent species from the monolignol without direct participation in radical coupling. If operative in lignifying tissues, such an effect could provide the organization necessary for replicating sequences of interunit linkages along macromolecular lignin chains. In this regard, the enzyme-catalysed dehydropolymerization of coniferyl alcohol would display the character of a typical template polymerization reaction. © 1997 Elsevier Science Ltd. All rights reserved

The use of organic markers in the differentiation of organic inputs to aquatic systems

In previous projects the estuarine distributions of a variety of molecular organic markers have been described and discussed in relation to sources, transport mechanisms and fates of anthropogenic and biogenic inputs to estuaries. Molecular markers have been used successfully to establish terrestrial inputs to marine water and to trace pollutants in water-ways. One of the components selected for study was lignin. Lignin compounds are phenolic polymers that occur as major constituents of the cell walls of vascular plants. Their source, natural abundance, wide distribution and resistance to microbial degradation render them good terrestrial markers and, via their phenolic aldehyde oxidation products, afford characterisation of their source material. In previous work, ratios of various lignin components suggest that permanently suspended material contains a significant proportion of degraded angiosperm tissues whereas, in resuspended material, a component of gymnosperm material is indicated. Comparison of the lignin concentrations in the suspended material with those in underlying sediment reveals that the permanently suspended material is preferentially enriched in lignin. This is due, at least in part, to the relative buoyancy of lignin-containing prticles which causes them to float in near-surface water. This paper considers whether such methodology can be usefully applied to the determination of terrestrial inputs to lentic environments.

THE COMPOSITION AND PHYLOGENETIC SIGNIFICANCE OF THE MOUGEOTIA (CHAROPHYCEAE) CELL WALL1

ABSTRACTThe two‐layered, fibrillar cell wall of Mougeotia C. Agardh sp. consisted of 63.6% non‐cellulosic carbohydrates and 13.4% cellulose. The orientation of cellulose microfibrils in the native cell wall agrees with the multinet growth hypothesis, which has been employed to explain the shift in microfibril orientation from transverse (inner wall) toward axial (outer wall). Monosaccharide analysis of isolated cell walls revealed the presence of ten sugars with glucose, xylose and galactose most abundant. Methylation analysis of the acid‐modified, 1 N NaOH insoluble residue fraction showed that it was composed almost exclusively of 4‐linked glucose, confirming the presence of cellulose. The major hemicellulosic carbohydrate was semi‐purified by DEAE Sephacel (Cl−) anion‐exchange chromatography of the hot 1 N NaOH soluble fraction. This hemicellulose was a xylan consisting of a 4‐xylosyl backbone and 2,4‐xylosyl branch points. The major hot water soluble neutral polysaccharide was identified as a 3‐linked galactan. Mougeotia cell wall composition is similar to that of (Charophyceae) and has homologies with vascular plant cell walls. Our observations support transtructural evidence which suggests that members of the Charophyceae represent the phylogenetic line that gave rise to vascular plants. Therefore, the primary cell walls of vascular plants many have evolved directly from structures typical of the filamentous green algal cell walls found in the Charophyceae.

Factors Controlling the Synthesis of Natural and Induced Lignins in Phleum and Elodea

Lignin is peculiar to the cell walls of vascular plants, and is produced only in specific cells at certain developmental periods. It is not detectable in aquatic vascular plants such as Elodea, but tisstues of these plants can be induced to form lignin-like products if suitable precursors are added (25). Since the products induced in Elodca were similar to those found naturally in other monocots such as Phleum, a comparison of the metabolism of phenolic compounds in these 2 organisms might aid in an understanding of the factors controlling lignification in plants. It is generally agreed that the phenolic monomers which are polymerized into lignin products arise from carbohydrates via the shikimic acid pathway (fig 1) (3). Some of the enzymes involved have been demonstrated in cell free extracts of higher plants (5). Mechanisms controlling lignification can be effective at any point of this synthetic pathway. In addition, since lignin is probably attached to specific sites in the wall, the number and type of these sites could be controlling factors. Possible regulatory agents of the terminal step of lignification have been studied in celery and peas by Siegel et al. (17,18) and in bamboo by Higuchi (10). The agents, indoleacetic acid (IAA), ascorbic acid and glutathione (GSH), were postulated to act as antioxidants. Possible regulatory mechanisms operating at an earlier level as well as at the peroxidation stage were investigated in this study of the metabolism of phenolic compounds associated with lignin biosynthesis in Elodea and Phleurn. In order to eliminate effects of treatments on growth bv elongation of the cell wall, nonelongating but still incompletely lignified tissues from these plants were chosen.