Ion exchange properties of plant root cell walls

Acid-base properties and the swelling capacity of wheat, lupin and pea root cell walls were investigated. Roots of seedlings and green plants of different age were analysed by the potentiometric method. The ion exchange capacity (S _i) and the swelling coefficient (K ^cw) of root cell walls were estimated at various pH values (from 2 to 12) and at different ionic strength (between 0.3 and 1000 mM). To analyse the polysigmoid titration curves pH_i = f (S _i), the Gregor's equation was employed. It was shown that the Gregor's model fits well the experimental data. The total number of the cation exchange (S _t ^cat) and the anion exchange (S _t ^an) groups were determined in the root cell walls. The number of the functional group of each type (S ^j) was estimated, and the corresponding values of pK _a ^j were calculated. It was shown that for all types of cation exchangeable groups arranged in the cell wall structure the acid properties are enhanced by the increasing concentration of electrolyte. For each ionogenic group the coefficients of Helfferich's equation [pK _a ^j = f (C ^K+)] were determined. It was found that the swelling of root cell walls changes with pH, C ^K+ and strongly depends on plant species. Within the experimental pH and C ^K+ range the swelling coefficient changes as follows: lupin > pea > wheat. The obtained results show that for the plant species under investigation the differences in the swelling coefficients originate from (a) the differences in the cross-linking degrees of polymeric chains arranged in the cell wall structure, (b) the differences in the number of carboxyl groups and (c) the differences in the total number of functional groups. Based on the estimated swelling coefficients in water it could be inferred that for wheat the cross-linking degree of the polymeric chains in the root cell walls is higher than those for lupin or pea. It has been emphasized that the calculated parameters (S ^j, pK _a ^j, K ^cw), the equation {pK _a ^j = f (C^K+)} and the dependencies {K ^cw = f (C^K+, pH)} allow to estimate quantitatively the changes in the ion exchange capacity of the root cell walls in response to the changes in an ionic composition of an outer solution. The results of these estimations allow to suggest that (a) the root apoplast is a compartment where the accumulation of cations takes place during the first stage of cation uptake from an outer medium, and (b) the accumulation degree is defined by pH and ionic composition of an outer solution. On the basis of the literature review and the results of the present experimental study it was proposed that the changes in the cell wall swelling in response to variances of environmental or experimental conditions could lead to a change of the water flow through a root apoplast. It has been supported that there is direct relationship between the swelling of root cell walls and the water flow within the plant root apoplast.     

Nonaqueous titration of amino groups in polymeric matrix of plant cell walls

Nonaqueous titration was used for detection of free amino groups in the polymeric matrix of plant cell walls. The content of amino groups varied in the range 0.54–0.91 and total nitrogen in the range 1.0–4.2 mmol per gram dry mass of cell walls depending on the plant species. However, these data on the high content of free amino groups do not correlate with the present day concept that the nitrogen fraction in charged amino groups in plant cell wall proteins, which are assumed to be mainly amino groups of lysine and arginine residues, is about 10%. It is supposed that most detected free amino groups belong to the hydroxy-amino acids hydroxyproline and tyrosine that can be bound at the hydroxyl group with the carbohydrate part of glycoprotein or another structural cell wall polymer.     

Some sulfonamide drugs inhibit ATPase activity of heat shock protein 90: investigation by docking simulation and experimental validation

Eight selected sulfonamide drugs were investigated as inhibitors of heat shock protein 90 (Hsp90). The investigation included simulated docking experiments to fit the selected compounds within the binding pocket of Hsp90. The selected molecules were found to readily fit within the ATP-binding pocket of Hsp90 in low-energy poses. The sulfonamides torsemide, sulfathiazole, and sulfadiazine were found to inhibit the ATPase activity of Hsp90 with IC₅₀ values of 1.0, 2.6, and 1.5 μM, respectively. Our results suggest that these well-established sulfonamides can be good leads for subsequent optimization into potent Hsp90 inhibitors.     

Accumulation of toxic metal ions on cell walls ofDatura innoxia suspension cell cultures

Summary Rapidly dividing cell suspension cultures derived fromDatura innoxia (Mill.) selectively remove certain toxic metal ions from nutrient and waste solutions. Many ions, including necessary micronutrients, bind tightly to different components of the primary cell wall. Cell viability is not required for metal chelation to the extracellular matrix, and biopolymers purified from these cultures can be used to selectively remove metal ions from waste streams. Binding of normally toxic metals to the primary cell wall significantly reduces their toxicity. Chemical and metal luminescence methods that generate information about metal binding and cell-wall components responsible for this are presented. The feasibility of using plant cells and their components for bioremediation is discussed. Presented in the Session-in-Depth Bioremediation through Biotechnological Means at the Congress on Cell and Tissue Culture, San Diego, CA, June 5–9, 1993.     

Ion-exchange properties of cell walls of Spinacia oleracea L. roots under different environmental salt conditions

Ion-exchange properties of the polymeric matrix of cell walls isolated from roots of 55-day-old Spinacia oleracea L. (Matador cv.) plants grown in nutrient solution in the presence of 0.5, 150, and 250 mM NaCl and from roots of Suaeda altissima L. Pall plants of the same age grown in the presence of 0.5 and 250 mM NaCl were studied. The ion-exchange capacity of the spinach cell walls was determined at pH values from 2 to 12 and different ionic strength of the solution (10 and 250 mM NaCl). In the structure of the root cell walls, four types of ionogenic groups were found: amine, two types of carboxyl (the first being galacturonic acid residue), and phenolic groups. The content of each type of group and their ionization constants were evaluated. The ion-exchange properties of spinach and the halophyte Suaeda altissima L. Pall were compared, and the qualitative composition of the ion-exchange groups in the cell walls of roots of these plants appeared to be the same and not depend on conditions of the root nutrition. The content of carboxyl groups of polygalacturonic acid changed in the cell walls of the glycophyte and halophyte depending on the salt concentration in the medium. These changes in the composition of functional groups of the cell wall polymers seemed to be a response of these plants to salt and were more pronounced in the halophyte. A sharp increase in the NaCl concentration in the medium caused a decrease in pH in the extracellular water space as a result of exchange reactions between sodium ions entering from the external solution and protons of carboxyl groups of the cell walls. The findings are discussed from the standpoint of involvement of root cell walls of different plant species in response to salinity.     

The One-Electron Reduction Potential of 3-Amino-1, 2, 4-benzotriazine 1, 4-dioxide (Tirapazamine): A Hypoxia-Selective Bioreductive Drug

The one-electron reduction potential of 3-amino-l, 2, 4-benzotriazine 1, 4-dioxide, tirapazamine (SR 4233) in aqueous solution has been determined by pulse radiol-ysis. Reversible electron transfer was achieved between radiolytically-generated one-electron reduced radicals of tirapazamine (T), and quinones or benzyl viologen as redox standards. The reduction potential E_m7(T/T^±) was -0.45 ± 0.01 V vs. NHE at pH 7. From the pH dependence of the reduction potential, pK_a = 5.6 ± 0.2 was estimated for the tirapazamine radical, a value similar to the pK_a determined by other methods.     

Calcium and the cell wall

Abstract. From this brief review it appears that the interactions between calcium ions and cell walls play a key role in plant physiology. Calcium ions are involved in many mechnisms: for example, stabilization of cell wall structures, acidic growth, ion exchange properties, control of the activities of wall enzymes. All these properties originate from the tight binding of calcium ions to the pectins present in the cell walls. The factor most important for controlling wall behaviour is the density of non-diffusible charges and, due to its high affinity, calcium can significantly affect this factor. We also discuss the theoretical ion exchange models in relation to the specific role of calcium ions.     

Studies of the chemical basis of the origin of protein synthesis: Initiation and direction of peptide growth

Summary The data presented in this paper show that the ease of non-enzymatic activation of carboxylic acids by ATP at pH 5 varies directly with the pK_a of the carboxyl group, and is consistent with the idea that it is the protonated form of the carboxyl group which participates in the activation reaction. Consequently, since most N-blocked amino acids have higher pK_a's than do their unblocked forms, they are activated more readily, and we have demonstrated that this principle applies to peptides as well,which are activated more rapidly than single amino acids. We propose that this fact may be partly responsible for the origin of two important features still observed in contemporary protein synthesis: (1) initiation in prokaryotes is accomplished with an N-blocked amino acid, and (2) elongation in all living systems occurs at the carboxyl end of the growing peptide.     

Immunocytochemical detection of lignin-related epitopes in cell walls in bryophytes and the charalean alga Nitella

Lignins are complex phenolic heteropolymers present in xylem and sclerenchyma cell walls in tracheophytes. The occurrence of lignin-like polymers in bryophytes is controversial. In this study two polyclonal antibodies against homoguaiacyl (G) and guaiacyl/syringyl (GS) synthetic lignin-like polymers that selectively labelled lignified cell walls in tracheophytes also bound to cell walls in bryophytes, the GS antibody usually giving a stronger labelling than the G antibody. In contrast to tracheophytes, the antibody binding in liverworts and mosses was not tissue-specific. In the hornworts Megaceros flagellaris and M. fuegiensis the pseudoelaters and spores were labelled more intensely than the other cell types with the GS antibody. The cell walls in Nitella were labelled with both antibodies but no binding was observed in Coleochaete. The results suggest that the ability to incorporate G or GS moieties in cell walls is a plesiomorphy (primitive character) of the land plant clade.     

Energetic analysis of the two controversial drug binding sites of the M2 proton channel in influenza A virus

Understanding the mechanism of the M2 proton channel of influenza A is crucially important to both basic research and drug discovery. Recently, the structure was determined independently by high-resolution NMR and X-ray crystallography. However, the two studies lead to completely different drug-binding mechanisms: the X-ray structure shows the drug blocking the pore from inside; whereas the NMR structure shows the drug inhibiting the channel from outside by an allosteric mechanism. Which one of the two is correct? To address this problem, we conducted an in-depth computational analysis. The conclusions drawn from various aspects, such as energetics, the channel-gating dynamic process, the pK_a shift and its impact on the channel, and the consistency with the previous functional studies, among others, are all in favour to the allosteric mechanism revealed by the NMR structure. The findings reported here may stimulate and encourage new strategies for developing effective drugs against influenza A, particularly in dealing with the drug-resistant problems.     

Taxonomic significance of cellulosic cell walls in the bangiales (Rhodophyta)

Cellulose has been characterized from isolated cell walls of the conchocelis phases of both Porphyra umbilicalis and P. leucostricta. Evidence for cellulose II (regenerated cellulose) in Schweitzer's reagent extracts was provided by X-ray powder analysis and paper chromatography of partial hydrolyzates. The presence of cellulose in the conchocelis phase of species of Porphyra provides evidence for the continuity of cell wall composition within the Rhodophyta. Adoption of a classification scheme incorporating consolidation of all red algal orders under the single class Rhodophyceae is proposed.     

Calcium ion transport across discs of the cortical flesh of apple fruit in relation to fruit development

Transport of Ca²⁺ through discs of apple fruit tissue was examined in tissue taken at different stages of fruit development. Transport rates decreased with fruit development when cation exchange was the predominant influence on transport (with 10⁻⁶ M ⁴⁵CaCl₂ as the source solution). This decrease was associated with a reduction in relative cell wall surface area, cation exchange capacity and cell wall yield that occurred during fruit growth. When diffusion was the major transport force, and when transport was influenced by solution infiltration of the tissue disc (10⁻² M ⁴⁵CaCl₂ in the source solution), transport rates increased during fruit growth. This increment was related to increases in air space of the tissue. Ca²⁺ transport through apple fruit tissue is influenced by the extent and nature of the cell wall, changing proportions of air space and Ca²⁺ concentration in the extracellular solution.     

Ion-Exchange Properties of Cell Walls Isolated from Lupine Roots

Acid–base properties of cell walls isolated from various root tissues of 7-day-old lupine seedlings and 14-day-old lupine plants grown in various media were studied. The ion-exchange capacity of root cell walls was estimated at various pH values (from 2 to 12) and constant ionic strength (10 mM). The parameters determining the qualitative and quantitative composition of cell wall ionogenic groups along the root length and in its radial direction were estimated using Gregor's model. This model fits the experimental data reasonably well. Four types of ionogenic groups were found in the cell walls: an amino group (pK _a 3), two types of carboxylic groups (pK _a 5 and 7.3, the first being the carboxylic group of galacturonic acid), and a phenolic group (pK _a 10). The number of functional groups of each type was estimated, and the corresponding ionization constant values were calculated. It is shown that the chemical composition of the ionogenic groups was constant along the root length as well as in its radial direction and did not depend on either physiological state or root nutrition, while the number of different groups varied. The content of carboxylic groups of -D-polygalacturonic acid in the root cell walls of 14-day-old plants was shown to depend on the distance from the root tip, being maximal in the zone of lateral roots. The number of these groups was 10- and 2-fold less in the central cylinder compared to that of cortex for 14-day-old plants and 7-day-old seedlings, respectively.     

Arrangement of mixed-linkage glucan and glucuronoarabinoxylan in the cell walls of growing maize roots

Background and Aims

Plant cell enlargement is unambiguously coupled to changes in cell wall architecture, and as such various studies have examined the modification of the proportions and structures of glucuronoarabinoxylan and mixed-linkage glucan in the course of cell elongation in grasses. However, there is still no clear understanding of the mutual arrangement of these matrix polymers with cellulose microfibrils and of the modification of this architecture during cell growth. This study aimed to determine the correspondence between the fine structure of grass cell walls and the course of the elongation process in roots of maize (Zea mays).

Methods

Enzymatic hydrolysis followed by biochemical analysis of derivatives was coupled with immunohistochemical detection of cell wall epitopes at different stages of cell development in a series of maize root zones.

Key Results

Two xylan-directed antibodies (LM11 and ABX) have distinct patterns of primary cell wall labelling in cross-sections of growing maize roots. The LM11 epitopes were masked by mixed-linkage glucan and were revealed only after lichenase treatment. They could be removed from the section by xylanase treatment. Accessibility of ABX epitopes was not affected by the lichenase treatment. Xylanase treatment released only part of the cell wall glucuronoarabinoxylan and produced two types of products: high-substituted (released in polymeric form) and low-substituted (released as low-molecular-mass fragments). The amount of the latter was highly correlated with the amount of mixed-linkage glucan.

Conclusions

Three domains of glucuronoarabinoxylan were determined: one separating cellulose microfibrils, one interacting with them and a middle domain between the two, which links them. The middle domain is masked by the mixed-linkage glucan. A model is proposed in which the mixed-linkage glucan serves as a gel-like filler of the space between the separating domain of the glucuronoarabinoxylan and the cellulose microfibrils. Space for glucan is provided along the middle domain, the proportion of which increases during cell elongation.     

Measurement of viscoelastic properties of root cell walls affected by low pH in lateral roots of Pisum sativum L.

Mechanical extensibility of the cell wall limits the elongation growth of roots. Low pH, ranging from pH 3–4.5, induces rapid elongation of excised roots, a phenomenon known as acid growth. The creep-extension analysis was carried out to measure and elucidate the viscoelastic properties of root cell walls in the acidic environment in vitro. The viscoelastic properties were determined at the elongation zone of the lateral roots of pea (Pisum sativumL. cv. Alaska) and described by the physical parameters of three elastic (E₀, E₁, E₂) and three viscosity (₀, ₁, ₂) parameters using a Kelvin–Voigt–Burgers' model. The present method could measure the viscoelasticity of 1-mm long root zones from 2 to 9 mm behind the tip. Among the parameters, E₀ and ₀ were the most significant parameters to represent the whole extensibility of the roots. The parameter ₀ markedly declined in response to the environmental low pH (acid growth), whereas other parameters were not much affected by low pH. Relationship between the change in these physical parameters and the change in cell wall extensibility under low pH was discussed in order to elucidate the rheological processes taking place in the elongating cell walls.     

Inducible expression of <Emphasis Type="Italic">Pisum sativum</Emphasis> xyloglucan fucosyltransferase in the pea root cap meristem,and effects of antisense mRNA expression on root cap cell wall structural integrity

Mitosis and cell wall synthesis in the legume root cap meristem can be induced and synchronized by the nondestructive removal of border cells from the cap periphery. Newly synthesized cells can be examined microscopically as they differentiate progressively during cap development, and ultimately detach as a new population of border cells. This system was used to demonstrate that Pisum sativum L. fucosyl transferase (PsFut1) mRNA expression is strongly expressed in root meristematic tissues, and is induced >2-fold during a 5-h period when mitosis in the root cap meristem is increased. Expression of PsFut1 antisense mRNA in pea hairy roots under the control of the CaMV35S promoter, which exhibits meristem localized expression in pea root caps, resulted in a 50-60% reduction in meristem localized endogenous PsFut1 mRNA expression measured using whole mount in situ hybridization. Changes in gross levels of cell wall fucosylated xyloglucan were not detected, but altered surface localization patterns were detected using whole mount immunolocalization with CCRC-M1, an antibody that recognizes fucosylated xyloglucan. Emerging hairy roots expressing antisense PsFut1 mRNA appeared normal macroscopically but scanning electron microscopy of tissues with altered CCRC-M1 localization patterns revealed wrinkled, collapsed cell surfaces. As individual border cells separated from the cap periphery, cell death occurred in correlation with extrusion of cellular contents through breaks in the wall.     

Chemical composition of the cell walls of Lolium multiflorum endosperm

Cell walls isolated from Lolium multiflorum endosperm grown in liquid suspension culture contain 90% carbohydrate (as anhydro-glucose), 0·3 nitrogen, 1·9% lipid and 4·3% ash. The relative proportions of neutral sugars present in hydrolysates of the wall polysaccharides are glucose, 50%; arabinose, 19%; xylose, 26% and galactose, 5%. Extraction of the wall with 7 M urea solubilizes a polysaccharide representing 19% of the wall and composed of glucose and minor amounts of pentoses. This fraction has been examined by acid and enzymic hydrolysis and by periodate oxidation, and was shown to be a β-1,3; 1,4-glucan with approx. 79% 1,4-linkages. A specific β-glucan hydrolase has been used to determine the content of this mixed-linked glucan in isolated endosperm cell walls.     

Celery (Apium graveolens L.) parenchyma cell walls examined by atomic force microscopy: effect of dehydration on cellulose microfibrils

Atomic force microscopy (AFM) was used to image celery (Apium graveolens L.) parenchyma cell walls in situ. Cellulose microfibrils could clearly be distinguished in topographic images of the cell wall. The microfibrils of the hydrated walls appeared smaller, more uniformly distributed, and less enmeshed than those of dried peels. In material that was kept hydrated at all times and imaged under water, the microfibril diameter was mainly in the range 6–25 nm. The cellulose microfibril diameters were highly dependent on the water content of the specimen. As the water content was decreased, by mixing ethanol with the bathing solution, the microfibril diameters increased. Upon complete dehydration of the specimen we observed a significant increase in microfibril diameter. The procedure used to dehydrate the parenchyma cells also influenced the size of cellulose microfibrils with freeze-dried material having larger diameters than air-dried material. Received: 16 November 1999 / Accepted: 7 March 2000