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D J Nevins

Publications and source records attributed to D J Nevins.

At least 19 recordsLinked to original sources
PubMed2000

New perspectives on the endo-beta-glucanases of glycosyl hydrolase Family 17.

Isozymes of glycosyl hydrolase Family 17 hydrolyze 1,3-beta-glucan polysaccharides found in the cell wall matrix of plants and fungi, enabling these plant enzymes to serve diverse roles in plant defense and plant development. Fourteen genes from Family 17 have been characterized in the genome of rice. A sequence dendrogram analysis divided these genes into four subfamilies. The recombinant GNS1 enzyme from subfamily B had 1,3;1,4-beta-glucanase activity, suggesting a role for this isozyme in plant development.

Amino Acid Sequence↗
PubMed2000

Endo-1,3;1,4-beta-glucanase from coleoptiles of rice and maize: role in the regulation of plant growth.

The Matrix Polymer Hydrolysis Model for regulation of growth in plants is based on the simultaneous hydrolysis and incorporation of new glucans into the cell wall observed in growing plant tissues. The inhibition of growth in rice coleoptile tissues treated with glucanase antibodies confirms similar results observed previously in maize coleoptiles and provides direct evidence for a role of glucanase in control of plant growth. Analysis of two-maize coleoptile endo-glucanase ESTs shows that these sequences are not related to any other previously known family of glycosyl hydrolase. Thus, the coleoptile endo-glucanase enzyme should be classified as a new enzyme group (E.C. 3.2.1.xx). These discoveries enable new initiatives for further investigation of the glucanase role in control of plant growth.

Base Sequence↗
PubMed2000

Cell wall autolytic activities and distribution of cell wall glucanases in Zea mays L. seedlings.

Exo- and endo-glucanases mediate specific degradation of cell wall (1,3)(1,4)-beta-D-glucans and these enzymes have been related to auxin-mediated growth and development of cereal coleoptiles. However, their distribution and functions have not been well established in other tissues. In this study the glucanase activities and cell wall autolytic activities of different maize organs were determined. Autolysis assays serve to evaluate the hydrolysis of cell wall polymers in situ by measuring the sugars released from the insoluble cell wall matrix resulting from the action of bound enzymes. Autolytic activities were observed in the cell walls of elongating young leaves, mesocotyl and roots of maize. Wall proteins extracted from all of these structures are enriched in several types of glucanases and other wall polysaccharide hydrolases. These enzymes therefore appear to have a widespread and fundamental role in wall metabolism in growing tissues.

Cell Wall↗
PubMed2000

Exo- and endoglucanases of maize coleoptile cell walls: their interaction and possible regulation.

Glucanase-mediated degradation of beta-(1,3)(1,4)-glucans has been attributed to auxin-induced cell wall loosening and thus tissue growth in cereal plants, but the regulatory mechanisms for the auxin-enhancement of glucanase activities in situ are not fully understood. Here, we report evidence for possible mechanisms which might account for auxin-induced changes in glucanase activities. A likely cause for acceleration of wall glucan degradation is the change in the ratio of exo- and endoglucanases. The combined enzymes synergistically promote beta-(1,3)(1,4)-glucan hydrolysis. In addition, these enzyme activities are enhanced by other enzymic and non-enzymic proteins and are also partially stimulated by divalent cations such as Ca(2+) and Mg(2+) at certain pH values. The acceleration of glucan degradation mediated by auxin may be mediated by changes and/or interactions of any of these factors in situ.

Cations↗
PubMed2000

Computer simulation of osmotic expansion and shrinkage in okra hypocotyl segments.

Okra hypocotyl segments were incubated in solutions of 0.3 or 0.4 M sorbitol at various temperatures and their shrinkage was measured. The result yielded an apparent activation energy for shrinkage of 4.8 kcal/mol, which is close to that of the viscosity of water. This coincidence suggests that the viscosity of water, i.e., the reciprocal function of water conductivity, is a limiting factor for osmotic shrinkage. Abrasion of okra hypocotyl segments with Carborundum substantially increased the rate of their osmotic shrinkage, indicating that the cuticle is the major barrier to water uptake by segments. The apparent activation energy for osmotic shrinkage was 4.5 kcal/mol in abraded segments. By introducing water conductivity into an algorithm, osmotic shrinkage and expansion of hypocotyl segments was successfully predicted by computation with this algorithm. Hence the extent of the contribution of water conductivity in osmotic shrinkage and expansion can be evaluated. Based on this simulation, water conductivity was identified as one of the major factors in governing the elongation growth rate of cells along with the osmotic pressure of the cell sap and the mechanical properties of the cell wall.

Cell Size↗
PubMed1997

Cell wall metabolism and autolytic activities of the yeast Saccharomyces exiguus.

Autolytic activities were measured in cell walls prepared from the yeast Saccharomyces exiguus. Walls of yeast cells exhibited higher autolytic activities directed toward glucans at the exponential phase of growth when compared to cells at the stationary phase, while glucanase activities in the soluble extract fraction were higher at the stationary phase when compared to exponential phase, suggesting an important role of cell wall glucanases in growth of the yeast cells. Yeast cell walls also exhibited a substantially high autolytic activity of glycoproteins containing mannose throughout growth. These results illustrate the diverse metabolism related to functions of yeast cell walls.

Carbohydrates↗
PubMed1996

Characterization of the isozymes of glucuronoxylan xylanohydrolase in the presence of a native cell wall substrate.

A simple but accurate method for measuring glucuronoxylan xylanohydrolase activity was developed using coleoptile cell wall particles prepared from maize (Zea mays L.). Two isozymes of glucuronoxylan xylanohydrolases designated as GX1 and GX2 (EC 3.2.1.136) were purified from a commercially available amylase preparation to electrophoretic homogeneity by three cation exchange chromatography steps. Upon characterization no significant differences between the two enzymes were detected: the molecular mass measured by MALDITOF mass spectrometry was 44,360 +/- 100 for GX1 and 44,370 +/- 50 for GX2 suggesting no difference in the total number of amino acid residues. Furthermore the N-terminal amino acid sequence for each of the isozymes was identical through the 37th amino acid residue. The values of pI were determined to be 9.0 for GX1 and 9.1 for GX2. The sensitivity to temperature, pH and to ionic strength was similar for both isozymes as were kinetic parameters including Km and Vmax. No differences could be detected in substrate specificity.

Amino Acid Sequence↗
PubMed1992

Structural characterization of an arabinoxylan-rhamnogalacturonan complex from cell walls of Zea shoots.

An arabinoxylan-rhamnogalacturonan complex, comprised of galacturonic acid, rhamnose, arabinose, xylose, and galactose in the ratios 75.9:4.6:5.2:3.5:5.4 and lesser amounts of other constituents, was dissociated from the water-insoluble matrix of cell walls of Zea mays by xylanase and glucuronoxylanase treatment. The solubilized complex retained its integrity when subjected to a series of separation procedures, and analysis of the sugar components throughout the elution profiles exhibited consistent ratios. The complex was subjected to controlled degradation by pectate lyase and pectin lyase, yielding two components comprised of rhamnose, fucose, arabinose, xylose, galactose, and galacturonic acid in the ratios 10.9:1.5:13.1:16.9:27.7:30.0 and 8.5:1.7:11.8:6.6:17.3:54.0, respectively, in addition to di-, tri-, and tetra-saccharides of galacturonic acid. The non-reducing terminals of the latter were characterized by the presence of 4,5-unsaturated hexuronic acid. The structural features of the two complex fractions were partially characterized.

Carbohydrate Sequence↗
PubMed1992

Comparison of the outer and inner epidermis : inhibition of auxin-induced elongation of maize coleoptiles by glucan antibodies.

Polyclonal antibodies, raised against beta-d-glucans prepared from oat (Avena sativa L.) caryopses, cross-reacted specifically with (1-->3),(1-->4)-beta-d-glucans when challenged in a dot blot analysis of related polymers bound to a cellulose thin layer chromatography plate. The antibodies suppressed indoleacetic acid (IAA)-induced elongation of segments from maize (Zea mays L.) coleoptiles when the outer surface was abraded. However, IAA-induced elongation of nonabraded segments or segments with abrasion restricted to the interior of the cylinder was not influenced by the antibodies. Fab fragments prepared from the antibodies gave similar results. The capacity for IAA to overcome outward curvature of split coleoptile segments was partially reversed by treatment of the segments with the antibodies. Fluorescence microscopy revealed that antibody penetration was largely restricted to the epidermal cell wall region. These results support the view that the degradation of (1-->3),(1-->4)-beta-d-glucans in the outer epidermal cell wall serves an essential role in auxin-induced elongation of Poaceae coleoptiles.

Journal Article↗
PubMed1991

Glucuronoxylan xylanohydrolase. A unique xylanase with the requirement for appendant glucuronosyl units.

A new category of beta-(1----4)-xylan xylanohydrolases that exhibit a specific capacity to hydrolyze glucuronoxylans was characterized using heteroxylans prepared from Vigna (Vigna angularis Ohwi et Ohashi cv. Takara) and maize (Zea mays L.) cell walls together with appropriate derivatives as substrates. Glucuronopyranosyl moieties, as side chains, were prerequisite for enzyme-mediated hydrolysis of the beta-(1----4)-xylosyl linkages. The enzyme degraded glucuronoxylans derived from Vigna cell walls to yield a major oligomeric species (formula; see text) where Xyl represents xylose and GlcA represents glucuronic acid. The enzyme also degraded glucuronoarabinoxylans derived from maize cell walls to yield a major oligomeric species containing a single glucuronosyl side chain and a single unsubstituted beta 1----4Xyl pendant terminal. These results indicate that this xylanohydrolase recognizes glucuronosyl moieties inserted as monomeric side chains along the xylan backbone and mediates the hydrolysis of the beta-(1----4)-xylosyl linkage of the adjacent unsubstituted xylosyl residue in heteroxylans. This enzyme is the first xylanohydrolase identified that recognizes distinctly different sugars constituting side chains. We propose to designate this new enzyme as a glucuronoxylan xylanohydrolase to be abbreviated as glucuronoxylanase. Use of this unique enzyme demonstrated the presence of repeating units in heteroxylans in cell walls of higher plants.

Bacillus subtilis↗
PubMed1991

Auxin-enhanced glucan autohydrolysis in maize coleoptile cell walls.

Cell walls isolated from auxin-pretreated maize (Zea mays L.) coleoptile segments were assayed to disclose evidence for the existence of enhanced autolysis. To improve the sensitivity of the measurements and to facilitate kinetic analysis, isolated cell walls were consolidated within a small column, and the autolysis rate was directly determined from the sugar content of the effluent. This protocol revealed that the maximum rate of autohydrolysis of walls prepared from segments occurs within the first 2 hours and a steady decline commences almost immediately. Walls from indoleacetic acid pretreated segments (0.5-4 hours) released sugar at a higher rate initially (110-125% of controls) and the enhanced rate of autolysis continued for 6 to 8 hours, but then it became equivalent to that of the controls. Pretreatment of the segments at acidic pH had no effect on the measurable rates of autolysis. The (1-->3), (1-->4)-beta-d-glucan content of the walls and the extractable glucanase activities support the hypothesis that temporal enhancement of autohydrolysis is a function of auxin on enzyme activity. The progressive decline in autolysis during prolonged incubations is consistent with the decrease in the quantity of the beta-d-glucan in the wall. The relationship between glucan content and autolysis rate is supported by the observation that while glucose pretreatment of segments had only a small effect on initial autolysis rates, the presence of the sugar during pretreatment served to extend the interval over which higher rates of autolysis could be sustained. The results demonstrate that autolysis is related to auxin-induced wall metabolism in maize coleoptiles.

Journal Article↗
PubMed1991

Inhibition of auxin-induced cell elongation of maize coleoptiles by antibodies specific for cell wall glucanases.

Polyclonal antibodies were raised in rabbits in response to the administration of purified exo- and endoglucanases extracted from cell walls of maize (Zea mays L. B37 x Mo17) coleoptiles. Since the antibodies formed specific conjugates when challenged with the glucanase antigens in immunoblot assays they were employed to evaluate the participation of glucanases in tissue growth. Indole-3-acetic acid induced cell elongation of abraded coleoptile segments was inhibited when the antibodies were supplied as a short term pretreatment (25-200 microgram/milliliter of serum protein). The extent of inhibition of IAA induced cell elongation was additive when endo- and exoglucanase antibodies were applied together. The results suggest that both enzymes have a role in mediating IAA-induced cell elongation. Pretreatment with exo- and endoglucanases antibodies also inhibited IAA induced degradation of noncellulosic beta-d-glucans and the increased level of cellulosic polymers in maize coleoptiles. Antibodies also inhibited the expression of the autohydrolytic degradation of glucans in isolated cell walls. The extent of inhibition was dependent on the antibody concentration applied. The results support the contention that enzymatic processes mediated by exo- and endoglucanases are responsible for cell wall autolytic reactions and that these reactions are linked to the mechanism for expressing auxin induced cell elongation in maize coleoptiles.

Journal Article↗
PubMed1991

Novel Technique for Measuring Tissue Firmness within Tomato (Lycopersicon esculentum Mill.) Fruit.

Developmental changes of tomato (Lycopersicon esculentum) fruit tissues during maturation were analyzed by a physically defined method (stress-relaxation analysis). The tip of a conical probe connected to a load sensor was positioned on the cut surface of a sliced tomato fruit, and the decay of the imposed stress was monitored. Stress-relaxation data thus obtained were used for the calculation of three stress-relaxation parameters. Different zones within tomato fruit harvested at six different ripening stages were analyzed. One of the stress-relaxation parameters, minimum stress-relaxation time (T(0)), decreased as the fruits matured. The decrease in T(0) was first found in the core of the carpel junction within the endopericarp at the blossom end during the breaker stage. The decrease in T(0) progressed from the blossom end, through the equatorial region and finally throughout the shoulder, as the fruit matured. In mature green fruit, T(0) values within the placenta and the proximal carpel junction were lower than those by other parts of the fruit. For all measurements the maximum stress-relaxation time was not substantially changed during maturation, nor were their changes observed in different regions of the fruit. The observed relaxation rate was therefore correlated with softening. The results indicate that fruit softening may be physically associated with the stress-relaxation parameter, T(0), and the extent of softening is a function of position within the fruit. Decreases in T(0) value appear to be correlated with the reported regional variation in the appearance of polygalacturonase.

Journal Article↗
PubMed1990

The Tomato Fruit Cell Wall : II. Polyuronide Metabolism in a Nonsoftening Tomato Mutant.

A nonsoftening tomato (Lycopersicon esculentum L.) variety, dg, was examined to assess the physiological basis for its inability to soften during ripening. Total uronic acid levels, 18 milligrams uronic acid/100 milligrams wall, and the extent of pectin esterification, 60 mole%, remained constant throughout fruit development in this mutant. The proportion of uronic acid susceptible to polygalacturonase in vitro also remained constant. Pretreatment of heat-inactivated dg fruit cell walls with tomato pectinmethylesterase enhances polygalacturonase susceptibility at all ripening stages. Pectinesterase activity of cell wall protein extracts from red ripe dg fruit was half that in extracts from analogous tissue of VF145B. Polygalacturonase activities of cell wall extracts, however, were similar in both varieties. Diffusion of uronic acid from tissue discs of both varieties increased beginning at the turning stage to a maximum of 2.0 milligrams uronic acid released/gram fresh weight at the ripe stage. The increased quantity of hydrolytic products released during ripening suggests the presence of in situ polygalacturonase activity. Low speed centrifugation was employed to induce efflux of uronide components from the cell wall tree space. In normal fruit, at the turning stage, 2.1 micrograms uronic acid/gram fresh weight was present in the eluant after 1 hour, and this value increased to a maximum of 8.2 micrograms uronic acid/gram fresh weight at the red ripe stage. However, centrifuge-aided extraction of hydrolytic products failed to provide evidence for in situ polygalacturonase activity in dg fruit. We conclude that pectinesterase and polygalacturonase enzymes are not active in situ during the ripening of dg fruit. This could account for the maintenance of firmness in ripe fruit tissue.

Journal Article↗
PubMed1990

Enzymic Analysis of Feruloylated Arabinoxylans (Feraxan) Derived from Zea mays Cell Walls : III. Structural Changes in the Feraxan during Coleoptile Elongation.

Changes in structural features of feraxan (feruloylated arabinoxylans) in cell walls during development of maize (Zea mays L.) coleoptiles were investigated by analysis of fragments released by feraxanase, a specific enzyme purified from Bacillus subtilis. The following patterns were identified: (a) The total quantity of carbohydrate dissociated from a given dry weight of cell wall by feraxanase remained rather constant throughout the initial 10 days of coleoptile development. However, during the same period the proportion of ferulic acid in the fraction increased 12-fold. The absolute amount of ferulic acid per coleoptile also increased rapidly during this developmental phase. (b) Fragments dissociated by the enzyme were resolved into feruloylated and nonferuloylated components by reversed phase chromatography. While the quantity of feruloylated fractions represented a small portion of the total arabinoxylan during the phase of maximum coleoptile elongation (days 2-4) this component increased in abundance to reach a plateau (after 8-10 days). In contrast, nonferuloylated fractions were most abundant during the stage of maximum elongation but declined to a constant level by day 6. (c) Glycosidic linkage analysis of carbohydrate reveals that substitution of the xylan backbone of feraxan by arabinosyl residues decreased during coleoptile growth. We conclude that significant incorporation of ferulic acid occurs and/or more feruloyated domains are added to the arabinoxylan during development. This augmentation in phenolic acids is accompanied by a concerted displacement of arabinosyl residues and/or a reduction in the incorporation of regions enriched in arabinosyl sidechains.

Journal Article↗
PubMed1989

Structure of a pectic polysaccharide fraction from zea shoots.

A pectic fraction, accounting for about 0.3% of the total cell wall polysaccharide, was derived from the hot water extract of an insoluble fraction of the buffer-homogenate of Zea shoots. The pectic polysaccharide fraction was characterized by fragmentation analysis after hydrolysis with acid and Erwinia carotovora pectate lyase. The results suggest that the fraction consists of mostly a linear homopolygalacturonan with neutral sugar components or a homogalacturonan and a rhamnogalacturonan with neutral sugar components.

Journal Article↗
PubMed1989

beta-d-Glucan Antibodies Inhibit Auxin-Induced Cell Elongation and Changes in the Cell Wall of Zea Coleoptile Segments.

Antiserum was raised against the Avena sativa L. caryopsis beta-d-glucan fraction with an average molecular weight of 1.5 x 10(4). Polyclonal antibodies recovered from the serum after Protein A-Sepharose column chromatography precipitated when cross-reacted with high molecular weight (1-->3), (1-->4)-beta-d-glucans. These antibodies were effective in suppression of cell wall autohydrolytic reactions and auxin-induced decreases in noncellulosic glucose content of the cell wall of maize (Zea mays L.) coleoptiles. The results indicate antibody-mediated interference with in situ beta-d-glucan degradation. The antibodies at a concentration of 200 micrograms per milliliter also suppress auxin-induced elongation by about 40% and cell wall loosening (measured by the minimum stress-relaxation time of the segments) of Zea coleoptiles. The suppression of elongation by antibodies was imposed without a lag period. Auxin-induced elongation, cell wall loosening, and chemical changes in the cell walls were near the levels of control tissues when segments were subjected to antibody preparation precipitated by a pretreatment with Avena caryopsis beta-d-glucans. These results support the idea that the degradation of (1-->3), (1-->4)-beta-d-glucans by cell wall enzymes is associated with the cell wall loosening responsible for auxin-induced elongation.

Journal Article↗