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PubMedDate not supplied

Reversible and strong immobilization of proteins by ionic exchange on supports coated with sulfate-dextran.

New and strong ionic exchange resins have been prepared by the simple and rapid ionic adsorption of anionic polymers (sulfate-dextran) on porous supports activated with the opposite ionic group (DEAE/MANAE). Ionic exchange properties of such composites were strongly dependent on the size of the ionic polymers as well as on the conditions of the ionic coating of the solids with the ionic polymers (optimal conditions were 400 mg of sulfate-dextran 5000 kDa per gram of support). Around 80% of the proteins contained in crude extracts from Escherichia coli and Acetobacter turbidans could be adsorbed on these porous composites even at pH 7. This interaction was stronger than that using conventional carboxymethyl cellulose (CMC) and even others such as supports coated with aspartic-dextran polymer. By means of the sequential use of the new supports and supports coated with polyethyleneimine (PEI), all proteins from crude extracts could be immobilized. In fact, a large percentage (over 50%) could be immobilized on both supports. Finally, some industrially relevant enzymes (beta-galactosidases from Aspergillus oryzae, Kluyveromyces lactis, and Thermussp. strain T2, lipases from Candida antarctica A and B, Candida rugosa, Rhizomucor miehei, and Rhyzopus oryzae and bovine pancreas trypsin and chymotrypsin) have been immobilized on these supports with very high activity recoveries and immobilization rates. After enzyme inactivation, the protein could be fully desorbed from the support, and then the support could be reused for several cycles. Moreover, in some instances the enzyme stability was significantly improved, mainly in the presence of organic solvents, perhaps as a consequence of the highly hydrophilic microenvironment of the support.

Adsorption↗
PubMed2001

The multicellular morphotypes of Salmonella typhimurium and Escherichia coli produce cellulose as the second component of the extracellular matrix.

Production of cellulose has been thought to be restricted to a few bacterial species such as the model organism Acetobacter xylinus. We show by enzymatic analysis and mass spectrometry that, besides thin aggregative fimbriae, the second component of the extracellular matrix of the multicellular morphotype (rdar) of Salmonella typhimurium and Escherichia coli is cellulose. The bcsA, bcsB, bcsZ and bcsC genes responsible for cellulose biosynthesis are not regulated by AgfD, the positive transcriptional regulator of the rdar morphotype. Transcription of the bcs genes was not co-expressed with the rdar morphotype under any of the environmental conditions examined. However, cellulose biosynthesis was turned on by the sole expression of adrA, a gene encoding a putative transmembrane protein regulated by agfD, indicating a novel pathway for the activation of cellulose synthesis. The co-expression of cellulose and thin aggregative fimbriae leads to the formation of a highly hydrophobic network with tightly packed cells aligned in parallel in a rigid matrix. As the production of cellulose would now appear to be a property widely distributed among bacteria, the function of the cellulose polymer in bacteria will have to be considered in a new light.

Arabidopsis Proteins↗
PubMed1999

In vitro synthesis and properties of pectin/Acetobacter xylinus cellulose composites

Pectin and cellulose are major components of most primary cell walls, yet little is known about the way in which they interact either during assembly or in subsequent functional performance of the wall. As a mimic of cell wall assembly, we studied the formation of molecular composites formed by deposition of cellulose from Acetobacter xylinus into pectin/calcium systems, and the molecular, architectural and mechanical properties of the composites obtained. The formation of interpenetrating cellulose/pectin composite networks (as envisaged in current models for primary cell walls) required a pre-existing, but not too strong, pectin network. For pectin either in solution or strongly networked, phase separation from cellulose occurred, providing two physical models for the formation of middle lamellae. Composite networks showed no evidence of direct molecular interaction between the components, but pectin networks became more aggregated following deposition of cellulose into them. The shear strength under small deformation conditions for cellulose/pectin composites was very similar to that of cellulose alone. In contrast, under uniaxial tension, extensibility was greatly increased and stiffness decreased. These major changes were due to the effect of pectin on cellulose network architecture at deposition, as they were maintained upon removal of the pectin component. These results show that the presence and physical state of pectin at the time of cellulose deposition in muro may be a significant determinant of subsequent extensibility without compromising strength.

Journal Article↗
PubMed1981

Enzymatic hydrolysis of cellulose: Visual characterization of the process.

Cellulose from the Gram-negative bacterium Acetobacter xylinum has been used as a model substrate for visualizing the action of cellulase enzymes from the fungus Trichoderma reesei. High-resolution electron microscopy reveals that A. xylinum normally produces a ribbon of cellulose that is a composite of bundles of crystalline microfibrils. Visual patterns of the process of cellulose degradation have been established. Enzymes are initially observed bound to the cellulose ribbon. Within 10 min, the ribbon is split along its long axis into bundles of microfibrils which are subsequently thinned until they are completely dissolved within 30 min. Incubations with purified components of the cellulase enzyme system produced less dramatic changes in ribbon structure. Purified 1,4-beta-D-glucan cellobiohydrolase I (D) (EC 3.2.1.91) produced no visible change in cellulose structure. Purified endo-1,4-beta-D-glucanase IV (EC 3.2.1.4) produced some splaying of ribbons into microfibril bundles. In both cases, whole ribbons were present even after 60 min of incubation, visually confirming the synergistic mode of action of these enzymes.

Journal Article↗
PubMed1991

Polypeptide composition of bacterial cyclic diguanylic acid-dependent cellulose synthase and the occurrence of immunologically crossreacting proteins in higher plants.

To comprehend the catalytic and regulatory mechanism of the cyclic diguanylic acid (c-di-GMP)-dependent cellulose synthase of Acetobacter xylinum and its relatedness to similar enzymes in other organisms, the structure of this enzyme was analyzed at the polypeptide level. The enzyme, purified 350-fold by enzyme-product entrapment, contains three major peptides (90, 67, and 54 kDa), which, based on direct photoaffinity and immunochemical labeling and amino acid sequence analysis, are constituents of the native cellulose synthase. Labeling of purified synthase with either [32P]c-di-GMP or [alpha-32P]UDP-glucose indicates that activator- and substrate-specific binding sites are most closely associated with the 67- and 54-kDa peptides, respectively, whereas marginal photolabeling is detected in the 90-kDa peptide. However, antibodies raised against a protein derived from the cellulose synthase structural gene (bcsB) specifically label all three peptides. Further, the N-terminal amino acid sequences determined for the 90- and 67-kDa peptides share a high degree of homology with the amino acid sequence deduced from the gene. We suggest that the structurally related 67- and 54-kDa peptides are fragments proteolytically derived from the 90-kDa peptide encoded by bcsB. The anti-cellulose synthase antibodies crossreact with a similar set of peptides derived from other cellulose-producing microorganisms and plants such as Agrobacterium tumefaciens, Rhizobium leguminosarum, mung bean, peas, barley, and cotton. The occurrence of such cellulose synthase-like structures in plant species suggests that a common enzymatic mechanism for cellulose biogenesis is employed throughout nature.

Affinity Labels↗
PubMed1992

Gene expression in cotton (Gossypium hirsutum L.) fiber: cloning of the mRNAs.

Cotton, an important natural fiber, is a differentiated epidermal cell. The number of genes that are active in fiber cells is similar to those in leaf, ovule, or root tissues. Through differential screening of a fiber cDNA library, we isolated five cDNA clones that are preferentially expressed in fiber. One of the cDNA clones, pCKE6, corresponded to an abundant mRNA in fiber. Transcripts for E6 were detected throughout the development of the fiber. Immunoprecipitation of in vitro translation products and Western blot analysis of fiber proteins showed two polypeptides in the range of 30-32 kDa as the products of E6 mRNA. Sequence analysis and hybrid-selected RNA translation also suggest that E6 mRNAs encode two polypeptides. Concentrations of E6 mRNA and protein are highest during the late primary cell wall and early secondary cell wall synthesis stages. Sequence comparison of E6 with other known eukaryotic and prokaryotic genes reveals no significant homology (GenBank; December 1991). E6 or a homologous gene(s) is conserved in several members of Malvaceae as well as in one other fiber-producing plant, kapok, but is not found in several other plants examined or in Acetobacter xylinum. A genomic clone corresponding to pCKE6 was isolated, and the promoter element of the E6 gene was shown to direct the expression of a carrot extensin mRNA in a tissue-specific and developmentally regulated fashion in transgenic cotton plants.

Amino Acid Sequence↗
PubMed1995

Molecular cloning of the isoquinoline 1-oxidoreductase genes from Pseudomonas diminuta 7, structural analysis of iorA and iorB, and sequence comparisons with other molybdenum-containing hydroxylases.

The iorA and iorB genes from the isoquinoline-degrading bacterium Pseudomonas diminuta 7, encoding the heterodimeric molybdo-iron-sulfur-protein isoquinoline 1-oxidoreductase, were cloned and sequenced. The deduced amino acid sequences IorA and IorB showed homologies (i) to the small (gamma) and large (alpha) subunits of complex molybdenum-containing hydroxylases (alpha beta gamma/alpha 2 beta 2 gamma 2) possessing a pterin molybdenum cofactor with a monooxo-monosulfido-type molybdenum center, (ii) to the N- and C-terminal regions of aldehyde oxidoreductase from Desulfovibrio gigas, and (iii) to the N- and C-terminal domains of eucaryotic xanthine dehydrogenases, respectively. The closest similarity to IorB was shown by aldehyde dehydrogenase (Adh) from the acetic acid bacterium Acetobacter polyoxogenes. Five conserved domains of IorB were identified by multiple sequence alignments. Whereas IorB and Adh showed an identical sequential arrangement of these conserved domains, in all other molybdenum-containing hydroxylases the relative position of "domain A" differed. IorA contained eight conserved cysteine residues. The amino acid pattern harboring the four cysteine residues proposed to ligate the Fe/S I cluster was homologous to the consensus binding site of bacterial and chloroplast-type [2Fe-2S] ferredoxins, whereas the pattern including the four cysteines assumed to ligate the Fe/S II center showed no similarities to any described [2Fe-2S] binding motif. The N-terminal region of IorB comprised a putative signal peptide similar to typical leader peptides, indicating that isoquinoline 1-oxidoreductase is associated with the cell membrane.

Amino Acid Sequence↗
PubMed1994

The solution structure of the circular trinucleotide cr(GpGpGp) determined by NMR and molecular mechanics calculation.

The 3'-5' circular trinucleotide cr(GpGpGp) was studied by means of 1D and 2D high resolution NMR techniques and molecular mechanics calculations. Analysis of the J-couplings, obtained from the 1H and 13C-NMR spectra, allowed the determination of the conformation of the sugar rings and of the 'circular' phosphate backbone. In the course of the investigations it was found that the Karplus-equation most recently parametrized for the CCOP J-coupling constants could not account for the measured J(C4'P) of 11.1 Hz and a new parametrization for both HCOP and CCOP coupling constants is therefore presented. Subsequent analysis of the coupling constants yielded 'fixed' values for the torsion angles beta and delta (with beta = 178 degrees and delta = 139 degrees). The value of the latter angle corresponds to an S-type sugar conformation. The torsion angles gamma and epsilon are involved in a rapid equilibrium in which they are converted between the gauche(+) and trans and between the trans and gauche(-) domain respectively. We show that the occurrence of epsilon in the gauche(-) domain necessitates S-type sugar conformations. Given the aforementioned values for beta, gamma, delta and epsilon the ring closure constraints for the ring, formed by the phosphate backbone can only be fulfilled if alpha and zeta adopt some special values. After energy minimization with the CHARMm force field only two combinations of alpha and zeta result in energetically favourable structures, i.e. the combination alpha (t)/zeta(g-) in case gamma is in a gauche(+) and epsilon is in a trans conformation, and the combination alpha (t)/zeta (g+) for the combination gamma (t)/epsilon (g-). The results are discussed in relation to earlier findings obtained for cd(ApAp) and cr(GpGp), the latter molecule being a regulator of the synthesis of cellulose in Acetobacter xylinum.

Guanine↗
PubMed1994

Overproduction and characterization of recombinant UDP-glucose pyrophosphorylase from Escherichia coli K-12.

Using oligonucleotide probes synthesized on the basis of partial amino acid sequences, we have cloned and sequenced the gene of Escherichia coli K-12 encoding UDP-glucose pyrophosphorylase. The gene consists of 906 base pairs and encodes a polypeptide of 302 amino acid residues with a calculated molecular weight of 32,941. Its nucleotide sequence was found to be identical with that recently registered (EMBL, X59940) for a gene coding for an unknown 33-kDa protein, which was later annotated as UDP-glucose pyrophosphorylase on the basis of genetic studies. The UDP-glucose pyrophosphorylase gene, mapped at 27.3 min in the E. coli chromosome, complemented the galU mutation, which renders the bacterium unable to ferment galactose. The recombinant enzyme overproduced in E. coli cells and purified to homogeneity catalyzed the synthesis and pyrophosphorolysis of UDP-glucose by a sequential mechanism. The enzyme required Mg2+ for maximal activity and was inhibited by free UTP and pyrophosphate. The E. coli enzyme shows significant sequence similarities with the enzymes from Acetobacter xylinum and Salmonella typhimurium. However, little or no similarity was found with the eukaryotic enzymes that are involved in the biosynthesis of storage carbohydrates, or with other enzymes acting on similar sugar nucleotides. Thus, UDP-glucose pyrophosphorylases participating in diverse metabolic pathways can be classified structurally into the prokaryotic and eukaryotic groups, even though they have almost identical catalytic properties.

Amino Acid Sequence↗
PubMed2000

Asaia bogorensis gen. nov., sp. nov., an unusual acetic acid bacterium in the alpha-Proteobacteria.

Eight Gram-negative, aerobic, rod-shaped and peritrichously flagellated strains were isolated from flowers of the orchid tree (Bauhinia purpurea) and of plumbago (Plumbago auriculata), and from fermented glutinous rice, all collected in Indonesia. The enrichment culture approach for acetic acid bacteria was employed, involving use of sorbitol medium at pH 3.5. All isolates grew well at pH 3.0 and 30 degrees C. They did not oxidize ethanol to acetic acid except for one strain that oxidized ethanol weakly, and 0.35% acetic acid inhibited their growth completely. However, they oxidized acetate and lactate to carbon dioxide and water. The isolates grew well on mannitol agar and on glutamate agar, and assimilated ammonium sulfate for growth on vitamin-free glucose medium. The isolates produced acid from D-glucose, D-fructose, L-sorbose, dulcitol and glycerol. The quinone system was Q-10. DNA base composition ranged from 59.3 to 61.0 mol% G + C. Studies of DNA relatedness showed that the isolates constitute a single species. Phylogenetic analysis based on their 16S rRNA gene sequences indicated that the isolates are located in the acetic acid bacteria lineage, but distant from the genera Acetobacter, Gluconobacter, Acidomonas and Gluconacetobacter. On the basis of the above characteristics, the name Asaia bogorensis gen. nov., sp. nov. is proposed for these isolates. The type strain is isolate 71T (= NRIC 0311T = JCM 10569T).

Acetic Acid↗
PubMed2001

Novel nitrogen-fixing acetic acid bacteria, Gluconacetobacter johannae sp. nov. and Gluconacetobacter azotocaptans sp. nov., associated with coffee plants.

Diazotrophic bacteria were isolated, in two different years, from the rhizosphere and rhizoplane of coffee (Coffea arabica L.) plants cultivated in Mexico; they were designated as type DOR and type SAd isolates. They showed characteristics of the family Acetobacteraceae, having some features in common with Gluconacetobacter (formerly Acetobacter) diazotrophicus, the only known N2-fixing species of the acetic acid bacteria, but they differed from this species with regard to several characteristics. Type DOR isolates can be differentiated phenotypically from type SAd isolates. Type DOR isolates and type SAd isolates can both be differentiated from Gluconacetobacter diazotrophicus by their growth features on culture media, their use of amino acids as nitrogen sources and their carbon-source usage. These results, together with the electrophoretic mobility patterns of metabolic enzymes and amplified rDNA restriction analysis, suggested that the type DOR and type SAd isolates represent two novel N2-fixing species. Comparative analysis of the 16S rRNA sequences revealed that strains CFN-Cf55T (type DOR isolate) and CFN-Ca54T (type SAd isolate) were closer to Gluconacetobacter diazotrophicus (both strains had sequence similarities of 98.3%) than to Gluconacetobacter liquefaciens, Gluconacetobacter sacchari (similarities < 98%) or any other acetobacteria. Strain CFN-Cf55T exhibited low levels of DNA-DNA reassociation with type SAd isolates (mean 42%) and strain CFN-Ca54T exhibited mean DNA-DNA reassociation of 39.5% with type DOR isolates. Strains CFN-Cf55T and CFN-Ca54T exhibited very low DNA reassociation levels, 7-21%, with other closely related acetobacterial species. On the basis of these results, two novel N2-fixing species are proposed for the family Acetobacteraceae, Gluconacetobacter johannae sp. nov. (for the type DOR isolates), with strain CFN-Cf55T (= ATCC 700987T = DSM 13595T) as the type strain, and Gluconacetobacter azotocaptans sp. nov. (for the type SAd isolates), with strain CFN-Ca54T (= ATCC 70098ST = DSM 13594T) as the type strain.

Acetic Acid↗
PubMed2000

Isolation of a novel insertion sequence from Mycobacterium fortuitum using a trap vector based on inactivation of a lacZ reporter gene.

An insertion sequence of Mycobacterium fortuitum has been isolated using a trap vector following insertion in and inactivation of the lacZ reporter gene. The trap vector is a temperature-sensitive (ts) Escherichia coli-mycobacterium shuttle plasmid, pCD4, which contains ts oriM, the kanamycin-resistance gene as a selection marker and a lacZ expression cassette. The ts mutation present in pCD4 functions in mycobacteria and enables screening for transposable elements from the mycobacterial genome that disrupt the lacZ gene by screening for white colonies on X-Gal plates in both mycobacterial as well as E. coli hosts. The vector was used to isolate a novel 1.653 kb insertion sequence from M. fortuitum named IS219. IS219 duplicated host DNA at the target site, had inverted repeats at its ends and contained two ORFs on one strand. One of the predicted proteins showed homology to a putative transposase from Acetobacter pasteurianus. IS219 was present in two copies in the genome of M. fortuitum. The trap vector appears to be useful in trapping insertion sequences from different mycobacteria by screening for the disrupted LacZ phenotype.

Amino Acid Sequence↗
PubMed2004

Swaminathania salitolerans gen. nov., sp. nov., a salt-tolerant, nitrogen-fixing and phosphate-solubilizing bacterium from wild rice (Porteresia coarctata Tateoka).

A novel species, Swaminathania salitolerans gen. nov., sp. nov., was isolated from the rhizosphere, roots and stems of salt-tolerant, mangrove-associated wild rice (Porteresia coarctata Tateoka) using nitrogen-free, semi-solid LGI medium at pH 5.5. Strains were Gram-negative, rod-shaped and motile with peritrichous flagella. The strains grew well in the presence of 0.35% acetic acid, 3% NaCl and 1% KNO3, and produced acid from l-arabinose, d-glucose, glycerol, ethanol, d-mannose, d-galactose and sorbitol. They oxidized ethanol and grew well on mannitol and glutamate agar. The fatty acids 18 : 1omega7c/omega9t/omega12t and 19 : 0cyclo omega8c constituted 30.41 and 11.80% total fatty acids, respectively, whereas 13 : 1 AT 12-13 was found at 0.53%. DNA G+C content was 57.6-59.9 mol% and the major quinone was Q-10. Phylogenetic analysis based on 16S rRNA gene sequences showed that these strains were related to the genera Acidomonas, Asaia, Acetobacter, Gluconacetobacter, Gluconobacter and Kozakia in the Acetobacteraceae. Isolates were able to fix nitrogen and solubilized phosphate in the presence of NaCl. Based on overall analysis of the tests and comparison with the characteristics of members of the Acetobacteraceae, a novel genus and species is proposed for these isolates, Swaminathania salitolerans gen. nov., sp. nov. The type strain is PA51T (=LMG 21291T=MTCC 3852T).

Acetic Acid↗
PubMed2004

Emendation of the genus Acidomonas Urakami, Tamaoka, Suzuki and Komagata 1989.

The genus Acidomonas and the species Acidomonas methanolica were recharacterized by using the type strain (NRIC 0498(T)), three reference strains and 10 methanol-utilizing bacteria that were isolated from activated sludge from three different sewage-treatment plants in Tokyo. Based on 16S rDNA sequences, all strains formed a single cluster within the Acetobacteraceae that was clearly different from the genera Acetobacter, Gluconobacter, Gluconacetobacter, Asaia and KOZAKIA: The 14 strains were identified as a single species, Acidomonas methanolica, by DNA-DNA similarities, showed DNA G+C contents that ranged from 62 to 63 mol% and had Q-10 as the major quinone, accounting for >87 % of total ubiquinones. Cells of Acidomonas methanolica had a single polar flagellum (or occasionally polar tuft flagella); this differs from a previous study that described peritrichous flagella. Oxidation of acetate was positive for all strains, but oxidation of lactate was weakly positive and varied with strains. Dihydroxyacetone was not produced from glycerol. Pantothenic acid was an essential requirement for growth. The strains tested grew at mostly the same extent at pH 3.0-8.0. Therefore, Acidomonas methanolica should be regarded as acidotolerant, not acidophilic. The descriptions of the genus Acidomonas and the species Acidomonas methanolica Urakami, Tamaoka, Suzuki and Komagata 1989 are emended with newly obtained data.

Acetobacteraceae↗
PubMed1994

A Nitrogen-Fixing Endophyte of Sugarcane Stems (A New Role for the Apoplast).

The intercellular spaces of sugarcane (Saccharum officinarum L.) stem parenchyma are filled with solution (determined by cryoscanning microscopy), which can be removed aseptically by centrifugation. It contained 12% sucrose (Suc; pH 5.5.) and yielded pure cultures of an acid-producing bacterium (approximately 104 bacteria/mL extracted fluid) on N-poor medium containing 10% Suc (pH 5.5). This bacterium was identical with the type culture of Acetobacter diazotrophicus, a recently discovered N2-fixing bacterium specific to sugarcane, with respect to nine biochemical and morphological characteristics, including acetylene reduction in air. Similar bacteria were observed in situ in the intercellular spaces. This demonstrates the presence of an N2-fixing endophyte living in apoplastic fluid of plant tissue and also that the fluid approximates the composition of the endophytes's optimal culture medium. The apoplastic fluid occupied 3% of the stem volume; this approximates 3 tons of fluid/ha of the crop. This endogenous culture broth consisting of substrate and N2-fixing bacteria may be enough volume to account for earlier reports that some cultivars of sugarcane are independent of N fertilizers. It is suggested that genetic manipulation of apoplastic fluid composition may facilitate the establishment of similar symbioses with endophytic bacteria in other crop plants.

Journal Article↗
PubMed1998

Bacterial cellulose-binding domain modulates in vitro elongation of different plant cells

Recombinant cellulose-binding domain (CBD) derived from the cellulolytic bacterium Clostridium cellulovorans was found to modulate the elongation of different plant cells in vitro. In peach (Prunus persica L.) pollen tubes, maximum elongation was observed at 50 &mgr;g mL-1 CBD. Pollen tube staining with calcofluor showed a loss of crystallinity in the tip zone of CBD-treated pollen tubes. At low concentrations CBD enhanced elongation of Arabidopsis roots. At high concentrations CBD dramatically inhibited root elongation in a dose-responsive manner. Maximum effect on root hair elongation was at 100 &mgr;g mL-1, whereas root elongation was inhibited at that concentration. CBD was found to compete with xyloglucan for binding to cellulose when CBD was added first to the cellulose, before the addition of xyloglucan. When Acetobacter xylinum L. was used as a model system, CBD was found to increase the rate of cellulose synthase in a dose-responsive manner, up to 5-fold compared with the control. Electron microscopy examination of the cellulose ribbons produced by A. xylinum showed that CBD treatment resulted in a splayed ribbon composed of separate fibrillar subunits, compared with a thin, uniform ribbon in the control.

Journal Article↗
PubMed1999

Roles of cellulose and xyloglucan in determining the mechanical properties of primary plant cell walls

The primary cell walls of growing and fleshy plant tissue mostly share a common set of molecular components, cellulose, xyloglucan (XyG), and pectin, that are required for both inherent strength and the ability to respond to cell expansion during growth. To probe molecular mechanisms underlying material properties, cell walls and analog composites from Acetobacter xylinus have been measured under small deformation and uniaxial extension conditions as a function of molecular composition. Small deformation oscillatory rheology shows a common frequency response for homogenized native cell walls, their sequential extraction residues, and bacterial cellulose alone. This behavior is characteristic of structuring via entanglement of cellulosic rods and is more important than cross-linking with XyG in determining shear moduli. Compared with cellulose alone, composites with XyG have lower stiffness and greater extensibility in uniaxial tension, despite being highly cross-linked at the molecular level. It is proposed that this is due to domains of cross-linked cellulose behaving as mechanical elements, whereas cellulose alone behaves as a mat of individual fibrils. The implication from this work is that XyG/cellulose networks provide a balance of extensibility and strength required by primary cell walls, which is not achievable with cellulose alone.

Journal Article↗
PubMed1986

Gel-Electrophoretic Separation, Detection, and Characterization of Plant and Bacterial UDP-Glucose Glucosyltransferases.

We have developed procedures for detection and characterization of UDP-glucose: glucosyltransferases following electrophoretic separation in nondenaturing polyacrylamide gels. Using digitonin-solubilized membrane protein preparations from a variety of plants and two cellulose-producing bacteria, activity can be demonstrated for several UDP-glucose:beta-glucan synthases with an in situ assay following gel electrophoresis. These enzymes can be characterized within the gels with respect to effector requirements and products produced, and several advantages of this assay over solution assays are demonstrated. For example, the clear dependence of plant UDP-glucose:(1-->3)-beta-glucan synthase on both Ca(2+) and a beta-linked glucoside is shown; bacterial cellulose synthases show direct stimulation within the gel by guanyl oligonucleotide, and the Acetobacter xylinum enzyme appears more stable in the gel assay than in solution assay.

Journal Article↗