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PubMed2003

Xylem-specific and tension stress-responsive expression of cellulose synthase genes from aspen trees.

Genetic improvement of cellulose biosynthesis in woody trees is one of the major goals of tree biotechnology research. Yet, progress in this field has been slow owing to (1) unavailability of key genes from tree genomes, (2) the inability to isolate active and intact cellulose synthase complexes and, (3) the limited understanding of the mechanistic processes involved in the wood cellulose development. Here I report on the recent advances in molecular genetics of cellulose synthases (CesA) from aspen trees. Two different types of cellulose synthases appear to be involved in cellulose deposition in primary and secondary walls in aspen xylem. The three distinct secondary CesAs from aspen- PtrCesA1, PtrCesA2, and PtrCesA3-appear to be aspen homologs of Arabidopsis secondary CesAs AtCesA8, AtCesA7, and AtCesA4, respectively, based on their high identity/similarity (>80%). These aspen CesA proteins share the transmembrane domain (TMD) structure that is typical of all known "true" CesA proteins: two TMDs toward the N-terminal and six TMDs toward the C-terminal. The putative catalytic domain is present between TMDs 2 and 3. All signature motifs of processive glycosyltransferases are also present in this catalytic domain. In a phylogenetic tree based on various predicted CesA proteins from Arabidopsis and aspen, aspen CesAs fall into families similar to those seen with Arabidopsis CesAs, suggesting their functional similarity. The coordinate expression of three aspen secondary CesAs in xylem and phloem fibers, along with their simultaneous tension stress-responsive upregulation, suggests that these three CesAs may play a pivotal role in biosynthesis of better-quality cellulose in secondary cell walls of plants. These results are likely to have a direct impact on genetic manipulation of trees in the future.

Arabidopsis↗

PubMed1993

Appearance of oxidized cellulose (Surgicel) on postoperative CT scans: similarity to postoperative abscess.

OBJECTIVE: Oxidized regenerated cellulose (Surgicel), a sterile knitted fabric that causes thrombus formation because of its physical properties, is frequently used for intraoperative hemostasis. Unlike traditional surgical sponges, it is bioabsorbable and can be left in the surgical bed. On CT scans, the appearance of the retained oxidized cellulose can mimic that of an abscess. The purpose of this study was to describe the appearance of oxidized regenerated cellulose on postoperative CT scans so that an erroneous diagnosis of an abscess can be avoided. MATERIALS AND METHODS: We reviewed the CT examinations of five postoperative patients in whom oxidized regenerated cellulose had been used for surgical hemostasis. Operative reports and surgeons confirmed the use of oxidized cellulose. Four CT scans were abdominal examinations, and one was a head examination. RESULTS: In four cases, CT scans showed focal, linear collections of gas within masses with mixed attenuation in or near the operative site. No air-fluid levels were present. In three patients, cultures of specimens obtained by aspiration were negative for pyogenic organisms. CONCLUSION: Retained oxidized cellulose can mimic an abscess on CT scans. Focal collections of air centrally located within a mass should alert the radiologist that oxidized cellulose may have been placed in the operative site, and an appropriate history should be sought.

Abscess↗

PubMed1992

Effect of soya and cellulose fibers on postprandial glycemic response in type II diabetic patients.

OBJECTIVE: We studied the effect on serum glucose and insulin of a preprandial ingestion of 7 g of soya fibers or of an equal amount of purified cellulose on eight non-insulin-dependent (type II) diabetic patients. RESEARCH DESIGN AND METHODS: Four tests were conducted in each patient in random order. In the first study, soya or cellulose was ingested before a standard breakfast, and postprandial glucose and insulin curves were determined. In the second study, intestinal absorption was investigated by means of a standard D-xylose absorption excretion test after the ingestion of soya or cellulose. RESULTS: Insulin profiles did not differ between the two treatments. The glycemic profiles after soya ingestion were lower than those after cellulose ingestion. The area under the glucose curve and glucose peaks were significantly higher after cellulose ingestion (area under the curve 20.2 +/- 3.88 vs. 15.57 +/- 4.42 mM x min, P = .05; glucose peaks 4.97 +/- 0.76 vs. 3.77 +/- 0.77 mM, P less than 0.02). The xylose tests were in the normal range, indicating that there was no interference with exose absorption, and no statistical difference was found between cellulose and soya treatment. CONCLUSIONS: It is concluded that soya fiber compared with purified cellulose has a favorable effect on the rise of postprandial glycemia in type II diabetic patients; moreover, the use of soya fibers did not carry any untoward side effect.

Blood Glucose↗

PubMed1997

Duraplasty with biosynthetic cellulose: an experimental study.

In the search for a new synthetic substitute for the dura mater, the authors conducted a research study using 32 mongrel dogs divided into three groups. Group I animals (21 dogs) underwent a right-sided parietooccipital craniotomy and substitution of two 1-cm pieces of dura mater by two different grafts: one piece of biosynthetic cellulose (50 mu thick) and one fragment of temporal fascia. The animals were observed for 30, 90, or 180 days. Group II animals (five dogs) underwent a somewhat larger craniotomy, removal of a 2-cm piece of dura mater, and lesioning of the cortex made by a thin sharp forceps, which caused bleeding that was controlled by application of a thin film of cellulose (10 mu thick). Duraplasty was performed using a 50-mu-thick cellulose membrane to complete the procedure and the animals were observed over a period of 270 days. Group III animals (six dogs) underwent smaller (1-cm diameter) bilateral parietal craniectomy, which included additional covering of the dura on the left side with 50-mu-thick cellulose and a suture of temporalis muscle. This group was observed for 40, 60, 80, or 120 days. Transient mild clinical symptoms were observed during the early postoperative period. At autopsy, macroscopic examination demonstrated good acceptance of the grafts with few and moderate extradural fibrosis, which caused adherence of the implants to the bone fragment. No adherence to the cortex was observed. Microscopic examination demonstrated absence of graft adherence to the cortical surface even when the cortex was injured. The cellulose was enveloped by two layers of connective tissue, the external layer being thicker than the internal one. Cellulose fibers increased in thickness over time until 30 days and then decreased in thickness until 270 days. This decrease in thickness between 30 to 270 days was statistically significant (p < 0.05). The physical properties of biosynthetic cellulose and the low cellular reaction to its implantation qualify this material as a dural substitute. Additional long-term studies must be undertaken to complete this report.

Animals↗

PubMed1980

The effects of dietary cellulose and sand on the combined metabolic plus endogenous energy and amino acid outputs of adult cockerels.

Two experiments were made with adult, Leghorn cockerels to measure the effects of inert materials on metabolic plus endogenous (M+E) excretions. In the first experiment birds starved for 24 hr were force-fed 25 g of either a laying hen diet (OHR) or corn supplemented with 0, 1, 2, 3, or 4 g of cellulose. Excreta voided were collected quantitatively during the two 24 hr periods postfeeding. A group of 4 birds was maintained unfed. Diets and excreta were assayed for amino acids (AA) and gross energy (GE). Supplemental cellulose had no effects on AA excretion nor on the availabilities of the AA within the OHR and corn. All AA, in the form of feed residues, were excreted within 24 hr of feeding. During the period 24 to 48 hr postfeeding the OHR fed birds excreted more AA than did the corn fed birds, probably because they were less depleted in protein. The AA excretions of the unfed birds decreased with the duration of starvation. The true available AA values for the diets were calculated using a series of three corrections for M+E AA outputs. The true metabolizable energy (TME) values of the OHR and corn were unchanged by the cellulose which indicates that the cellulose contained no available energy and had no effect on the availability of the energy within the OHR and corn. It also demonstrated that the cellulose did not change the M+E energy excretion. The second experiment was a 4 X 4 factorial with four diets: nil, corn, wheat, oats (25 g/bird), and four levels of sand: 0, 1, 2, 3 g/bird. The birds were starved for 24 hr prior to feeding and excreta were collected quantitatively during the two 24 hr periods postfeeding. Feeding sand to the birds had no effect on their GE outputs but it was noted that little sand was voided within 48 hr postfeeding. The TME values were determined for the grains. A number of significant differences were observed but there was no evidence that the sand altered the TME values. It was therefore concluded that the sand had no effect on the M+E energy excretion of the assay birds. It is concluded that supplemental cellulose had no effect on the AA excretion of adult Leghorn cockerels. Neither cellulose nor sand, in the levels fed, altered M+E energy excretion. Neither diluent appeared to alter the utilization of the materials with which they were fed.

Amino Acids↗

PubMed1999

Digestion of crystalline cellulose substrates by the clostridium thermocellum cellulosome: structural and morphological aspects.

The action of cellulosomes from Clostridium thermocellum on model cellulose microfibrils from Acetobacter xylinum and cellulose microcrystals from Valonia ventricosa was investigated. The biodegradation of these substrates was followed by transmission electron microscopy, Fourier-transform IR spectroscopy and X-ray diffraction analysis, as a function of the extent of degradation. The cellulosomes were very effective in catalysing the complete digestion of bacterial cellulose, but the total degradation of Valonia microcrystals was achieved more slowly. Ultrastructural observations during the digestion process suggested that the rapid degradation of bacterial cellulose was the result of a very efficient synergistic action of the various enzymic components that are attached to the scaffolding protein of the cellulosomes. The degraded Valonia sample assumed various shapes, ranging from thinned-down microcrystals to crystals where one end was pointed and the other intact. This complexity may be correlated with the multi-enzyme content of the cellulosomes and possibly to a diversity of the cellulosome composition within a given batch. Another aspect of the digestion of model celluloses by cellulosomes is the relative invariability of their crystallinity, together with their Ialpha/Ibeta composition throughout the degradation process. Comparison of the action of cellulosomes with that of fungal enzymes indicated that the degradation of cellulose crystals by cellulosomes occurred with only limited levels of processivity, in contrast with the observations reported for fungal enzymes. The findings were consistent with a mechanism whereby initial attack by a cellulosome of an individual cellulose crystal results in its 'commitment' towards complete degradation.

Acetobacter↗

PubMed1992

Gravity effects on cellulose assembly.

The effect of microgravity on cellulose synthesis using the model system of Acetobacter xylinum was the subject of recent investigations using The National Aeronautics and Space Administration's Reduced Gravity Laboratory, a modified KC-135 aircraft designed to produce 20 sec of microgravity during the top of a parabolic dive. Approximately 40 parabolas were executed per mission, and a period of 2 x g was integral to the pullout phase of each parabola. Cellulose biosynthesis was initiated on agar surfaces, liquid growth medium, and buffered glucose during parabolic flight and terminated with 2.0% sodium azide or 50.0% ethanol. While careful ground and in-flight controls indicated normal, compact ribbons of microbial cellulose, data from five different flights consistently showed that during progression into the parabola regime, the cellulose ribbons became splayed. This observation suggests that some element of the parabola (the 20 sec microgravity phase, the 20 sec 2 x g phase, or a combination of both) was responsible for this effect. Presumably the cellulose I alpha crystalline polymorph normally is produced under strain, and the microgravity/hypergravity combination may relieve this stress to produce splayed ribbons. An in-flight video microscopy analysis of bacterial motions during a parabolic series demonstrated that the bacteria continue to synthesize cellulose during all phases of the parabolic series. Thus, the splaying may be a reflection of a more subtle alteration such as reduction of intermicrofibrillar hydrogen bonding. Long-term microgravity exposures during spaceflight will be necessary to fully understand the cellulose alterations from the short-term microgravity experiments.

Acetobacter↗

PubMed2004

[Application of high-efficient cellulose utilization microorganisms in co-composting of vegetable wastes and flower stalk].

An inoculation composing 17 species of cellulose utilization microorganisms was used in co-composting of vegetable wastes and flower stalk, and the efficiency of the inoculation on lignocellulose degradation was studied. The experiments result show that at the beginning of the first stage of composting, inoculating cellulose utilization microorganism in the substrates with 0.5% (V/V) can improve the biomass of the microorganisms into the substrates greatly and make them dominant ones. When the temperature was controlled as 55 degrees C, the biomass of cellulose utilization microorganisms can keep between [symbol: see text] 3.84 x 10(9)-1.80 x 10(10) CFU/g. At the beginning of the second stage, inoculating with 1% (V/V) can improve the temperature during this period effectively. Monitoring of the content of lignocelluloses in the substrate shows that the inoculation of cellulose utilization microorganism can accelerate the degradation of cellulose. The degradation efficiency of cellulose under inoculation condition is 23.64% higher than those without inoculation. This shows that inoculating with cellulose utilization microorganisms in each stage of the composting can greatly decompose the lignocellulose in the substrates, accelerate the co-composting process and improve the quality of composting production.

Bacteria↗

PubMed2005

Molecular basis of cellulose biosynthesis disappearance in submerged culture of Acetobacter xylinum.

Acetobacter xylinum strains are known as very efficient producers of bacterial cellulose which, due to its unique properties, has great application potential. One of the most important problems faced during cellulose synthesis by these bacteria is generation of cellulose non-producing cells, which can appear under submerged culture conditions. The reasons of this remain unknown. These studies have been undertaken to compare at the molecular level wild-type, cellulose producing (Cel(+)) A. xylinum strains with Cel(-) forms of cellulose-negative phenotype. Comparison of protein profiles of both forms of A. xylinum by 2D electrophoresis allowed for the isolation of proteins which were produced exclusively by either Cel+ or Cel- cells. Sequences of peptides derived from these proteins were aligned with those of proteins deposited in databases. This analysis revealed that Cel(-) cells lacked two enzymes: phosphoglucomutase and glucose-1-phosphate uridylyltransferase, which generates UDP-glucose being the substrate for cellulose synthase. DNA was analyzed by ligation-mediated PCR carried out at low denaturation temperature (PCR-MP). Two DNA fragments of different thermal stability (218 and 217 bp) were obtained from the DNA of Cel(+) and Cel(-) forms, respectively. The only difference between these Cel(-) and Cel(+) DNA fragments is deletion of one T residue. Alignment of those two sequences with those deposited in the GenBank database revealed that similar fragments are present in the genomes of some bacterial cellulose producers and are located downstream from open reading frames (ORF) encoding phosphoglucomutase. The meaning of this observation is discussed.

Base Sequence↗

PubMed1989

Biologically active polymer supports based on cellulosic derivatives: synthesis and kinetic study.

Bioactive cellulose derivatives have been synthesised by coupling enzymes/antibiotics on carboxymethyl cellulose acid chloride and cellulose carbonate. The effect of pH and temperature on the enzymatic activity of amyloglucosidase immobilised on cellulose carbonate was studied. Michaelis-Menten kinetics have been obeyed to the first degree of approximation despite the restricted mobility of the attached enzyme on the polymer support. Lineweaver-Burk plots for the amyloglucosidase immobilized on carboxymethyl cellulose acid chloride at ambient pH with cellulose carbonate at pH 8 have also been plotted. The Michaelis-Menten constant for the immobilized amyloglucosidase on cellulose carbonate at pH 8 was 9.1 mM, and the activation energy for starch hydrolysis was 21.8 kcals/mole.

Cellulose↗

PubMed1989

Proliferative activity in the colon of the mouse and its modulation by dietary starch, fat, and cellulose.

The effect of dietary starch, fat, and cellulose on colonic proliferation was studied in female C57Bl/6J mice after 4 weeks of feeding with diets containing various levels of starch (3, 36, and 57-65%), various levels of fat (corn oil, 5 and 29%) and various levels of cellulose (2 and 10%). Cell proliferation was measured by colchicine arrest and [3H]thymidine incorporation. The following parameters were analyzed: mitotic index, labeling index, and position of labelled cells along the crypt. Increasing starch content from 3 to 36% decreased cell proliferation both in low (5%) and high (29%) fat diets. By estimating simultaneously the effects of starch and fat with a single multiple regression model, we observed a decrease of mitotic index from 3.04 +/- 0.34 to 2.04 +/- 0.43 (means +/- SE) (P less than 0.05) when starch was increased from 3 to 36% regardless of the level of fat. Other proliferation parameters showed a similar pattern. Changes in dietary fat alone did not affect significantly cell proliferation. We also investigated the effect of starch at high levels (57-65%) and its interactions with cellulose. High starch (57-65%) increased the labeling index from 7.70 +/- 0.58 to 9.65 +/- 0.88 (P less than 0.05), when also considering the effect of cellulose in the multiple regression model. Cellulose by itself did not change the labeling index. Varying starch from 36 to 57-65% increased the number of cells/crypt column from 22.20 +/- 0.82 to 25.87 +/- 1.21 (P less than 0.05) and varying cellulose from 2 to 10% increased the number of cells/crypt column from 22.20 +/- 0.82 to 27.25 +/- 0.97 (P less than 0.01). The results indicate that either high or low fat diets, containing 36% starch, have the minimum proliferative effects in the mouse colon. However, diets containing high levels of both starch (57%) and cellulose (10%) may induce an increase in proliferation. These data suggest a potential beneficial effect of starch on colon proliferation.

Animals↗

PubMed1997

Transforming growth factor alpha distribution in rectal crypts as a biomarker of decreased colon cancer risk in patients consuming cellulose.

Data from rat experimental carcinogenesis studies indicate that supplemental dietary cellulose reduces the incidence of colon cancer. Epidemiology studies also indicate that high dietary fiber reduces the risk of colorectal cancer in humans. Patients diagnosed with sporadic adenomas were entered into a randomized clinical trial to determine if supplemental dietary cellulose would reduce the patients' risk for colon cancer. Immunohistochemical staining for transforming growth factor alpha (TGF-alpha) was done on biopsies of rectal mucosa taken from patients at the time of initial polypectomy and 1 year later. Results were evaluated for utility as a surrogate end point biomarker for reduction in colon cancer risk. There was a significant decrease in the fraction of the rectal crypt cells that stained for TGF-alpha in six of seven of the patients given the cellulose supplements but in only one of six of the patients not given cellulose. Thus, whether evaluated as a group or in individual patients, there was a significant decrease in TGF-alpha in rectal crypts due to cellulose intervention, which correlated with the expected ability of supplemental dietary cellulose to decrease the risk for colon cancer. Long-term testing of the ability of dietary cellulose to reduce adenoma recurrence is under way to validate the use of TGF-alpha as a surrogate end point biomarker.

Adult↗

PubMed2006

Rapid online equilibration method to determine the D/H ratios of non-exchangeable hydrogen in cellulose.

An improved method for the determination of deuterium-to-hydrogen (D/H) ratios of non-exchangeable hydrogen in cellulose is presented. The method is based on the equilibration reaction of the hydroxyl hydrogen of cellulose and water vapour of known isotopic composition. The equilibrated cellulose is pyrolysed and the total D/H ratio determined by subsequent online isotope ratio mass spectrometry (IRMS). With a mass balance system the D/H ratio of non-exchangeable hydrogen is recalculated after an empirical calibration has been performed, yielding a mean exchangeability of 0.239 and an equilibrium fractionation factor of 1.082 between the hydroxyl hydrogen of cellulose and water hydrogen at 110 degrees C. Equilibration takes 10 min per sample. Results obtained by this online equilibration method agree very well with values obtained by the nitration technique (R2 = 0.941). The uncertainty of the equilibration method is +/-4 per thousand resulting from a single standard deviation of +/-2.8 per thousand for the equilibration determined by standard cellulose and 2.8 per thousand from the variable exchangeability of the hydroxyl hydrogen in cellulose due to crystalline areas. The latter uncertainty may be lowered by minimising the crystallinity of the cellulose. Advantages of this new technique are (i) the considerably reduced sample amount required (as low as 0.2 mg, ideally 0.5 mg compared with 20 mg for the conventional nitration technique); (ii) an approximately 100-fold reduced process time; and (iii) no need for the hazardous chemicals used in the nitration technique.

Journal Article↗

PubMed2006

Molecularly smooth cellulose surfaces for adhesion studies.

Cellulose is deposited on silicon wafer surfaces via spin coating from a solution of cellulose in dimethylacetamide (+7% lithium chloride). The resulting cellulose layers were analyzed by ellipsometry, AFM, FTIR, ICP-MS, X-ray reflectivity, and contact angle measurements. For cellulose concentrations below 0.07 wt% the wafer surfaces are covered with a network of cellulose fibrils. For concentrations between 0.07 and 0.5 wt%, closed films with thicknesses between 1.5 and approximately 10 nm are obtained. These films are molecularly smooth (rms roughness<2 nm). Higher concentrations result in thicker films with significantly rougher surfaces (rms roughness>2 nm). The cellulose layers were used to investigate cellulose/cellulose adhesion and their modification by polyelectrolytes. To this end the sticking behavior of cellulose beads was analyzed. It is demonstrated that the sticking of the beads depends on the type, amount, and adsorption symmetry of adsorbed polyelectrolyte. Low, incomplete polyelectrolyte coverage always enhances sticking, whereas for high coverage the symmetry of the polyelectrolyte coating is very important. In this case, sticking (adhesion) is enhanced if only one surface is covered with polyelectrolyte prior to contact. If both surfaces were fully covered with polyelectrolytes before contact, sticking (adhesion) is decreased.

Journal Article↗

PubMed2005

Influence of ultrasound treatment on accessibility and regioselective oxidation reactivity of cellulose.

Cellulose fibers were treated with ultrasound in order to improve the accessibility and the reactivity of cellulose. The influence of ultrasound treatment on changes of morphology structure, accessibility and oxidation reactivity of cellulose with sodium periodate were discussed. The results revealed an increase in cellulose's accessibility in terms of water retention value (WRV) with increasing ultrasound treatment time, corresponding to 73.0%, 75.6%, 80.8%, 98.7% and 119.0% after treated for 0, 90, 180, 360 and 720 s, respectively. Furthermore, the regioselective oxidation reactivity of cellulose with sodium periodate was also successfully improved by the ultrasound treatment. However, no significant changes in crystallinity of cellulose were noted after ultrasound treatment. The oxidized products dialdehyde cellulose (DAC) was further characterized by means of FTIR, X-ray diffraction and SEM.

Journal Article↗

PubMed1997

Ex vivo complement protein adsorption on positively and negatively charged cellulose dialyser membranes.

An ex vivo test system was used to measure complement protein C3 and factor B adsorption onto small dialyser modules made from regenerated and modified cellulosic hollow fibre membranes in which positive diethylaminoethyl (DEAE) or negative carboxymethyl (CM) groups were introduced into the cellulose matrix. The extracorporeal system, which included test-dialysers and the dialysis environment, allowed the use of labelled proteins without contaminating the blood donors which were connected in an open-loop fashion to the extracorporeal test system. The modules were removed at selected time points from the extracorporeal system for radioactivity counting. The results were used to evaluate the mechanisms involved in complement reactions to foreign surfaces. The system therefore allowed the analysis of complement protein adsorption occurring in the dialyser modules and its relationship to the complement generation rate in the extracorporeal system to be evaluated. It was possible to demonstrate that significant complement C3 and factor B adsorption occurred in the test modules made of cellulosic membranes. Complement adsorption as a function of the pH and the release reaction of the adsorbed C3 and factor B after membrane blood perfusion were therefore found to be variable according to the cellulosic membrane type and the presence of positive or negative charged groups within the cellulose matrix. The data obtained from the ex vivo model therefore provided additional evidence on the discussion of the mechanisms involved in the increased complement activation by regenerated cellulose and in its attenuation by DEAE- or CM-modified cellulose.

Journal Article↗

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↗

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↗