Production of cellulose from glucose by Acetobacter xylinum

Acetobacter xylinum 1FO 13693 was selected as the best cellulose-producing bacterium among 41 strains belonging to the genus Acetobacter and Agrobacterium. Cellulose was found to be produced at the liquid surface in static liquid cultivation. The rate of cellulose production depended proportionally on the surface-area of the culture medium and was unaffected by the depth and volume of the medium. The optimum pH for cellulose production was 4.0 to 6.0. Glucose, fructose and glycerol were preferred carbon sources for cellulose production. The yield of cellulose, relative to the glucose consumed, decreased with an increase in initial glucose concentration, and gluconic acid accumulated at a high initial glucose concentration. The decrease in cellulose yield could be due to some glucose being metabolized to gluconic acid. However, the accumulated gluconic acid did not affect cellulose production. The culture conditions of the bacterium for cellulose production were optimized. The maximum production rate of cellulose was 36 g/d·m², with a yield of 100% for added glucose under the optimal conditions.     

Effects of acetan on production of bacterial cellulose by Acetobacter xylinum

Acetan is a water-soluble polysaccharide produced by a bacterial cellulose (BC) producer, Acetobacter xylinum. An acetan-nonproducing mutant, EP1, was generated from wild-type A. xylinum BPR2001 by the disruption of aceA, which may act to catalyze the first step of the acetan biosynthetic pathway in this bacterium. EP1 produced less BC than the wild-type strain. However, when EP1 was cultured in a medium containing acetan, BC production was stimulated and the final yield of BC was equivalent to that of BPR2001. The culture broth containing acetan was more viscous and the free cell number was higher than that of the broth without the polysaccharide, so acetan may hinder the coagulation of BC in the broth. The addition of 1.5 g/l agar also increased BC production; we concluded that acetan and BC syntheses were not directly related on the genetic level.     

Induction of orientation of bacterial cellulose microfibrils by a novel terpenoid from Acetobacter xylinum

1. The bacterium Acetobacter xylinum produces extracellular cellulose microfibrils that form a pellicle in the medium enmeshing the bacterial cells. These microfibrils may show some localized alignment, which can be seen as birefringence when the culture is viewed between crossed Polaroid sheets. 2. An increase in birefringence can be induced by the addition of small amounts of certain classes of lipids, particularly sterols, to the cultures. 3. A crude lipid extract from Acetobacter cells induced greatly increased birefringence when added to fresh cultures of this organism. 4. When the bacterial lipids were fractionated, most of the activity was recovered in a complex, polar lipid. The lipid is secreted into the medium during growth and is unstable. The non-saponifiable portion of this lipid is shown to be a 1:1 mixture of a saturated and a monounsaturated C₃₅ tetrahydroxy terpene with a hopane ring system in the accompanying paper by Förster et al. (1973). The saturated molecule is referred to as tetrahydroxybacteriohopane. 5. Tetrahydroxybacteriohopane is itself capable of inducing birefringence in cultures as is 22-hydroxyhopane, which was also isolated from the non-saponifiable fraction of the total lipids. 6. The mechanism of induction of birefringence (orientation of microfibrils) is not known. This is unlikely to be a specific effect, since all the above compounds are active (intact lipid, tetrahydroxybacteriohopane, 22-hydroxyhopane), as are other classes of lipid. It is suggested, however, that a common mechanism may be involved and that similar compounds may be concerned with control of microfibril alignment in the cells of higher plants.     

Relationship between Sulfaguanidine Resistance and Increased Cellulose Production in Acetobacter xylinum BPR3001E

The mechanism of the increased cell growth and cellulose production of Acetobacter xylinum subsp. sucrofermentans BPR3001E, a sulfaguanidine (SG)-resistant mutant, was investigated. We found that adding p-aminobenzoic acid (PABA) to cultures of the parent strain, BPR2001, led to increased levels of intracellular adenosine-related purine compounds and increased cellulose production. Furthermore, adding ATP increased the cellulose production by permeabilized BPR2001 cells. On the other hand, the intracellular levels of PABA and adenosine-related purine compounds in BPR3001E cells were higher than those in BPR2001 cells. These results suggest that SG resistance increases enhance cellulose production through increased levels of intracellular high-energy compounds caused by increased PABA biosynthesis, reflecting the promoted supply of cellulose precursors.     

Utilization of the buffering capacity of corn steep liquor in bacterial cellulose production by Acetobacter xylinum

Acetobacter xylinum BPR2001 produces water-insoluble bacterial cellulose (BC). Using a pH sensor for the accurate control of pH, which is one of the most critical factors for efficient BC production, is difficult especially in a baffled shake-flask and an airlift reactor. The buffering capacity of corn steep liquor (CSL) was estimated by measuring (buffering capacity) values in advance and was used to maintain the pH within the optimal range during the production of BC. When CSL was added to either a shake-flask, a stirred-tank reactor or an airlift reactor, BC production was almost the same as that in cultivations where pH was controlled manually or by a pH sensor.     

Production of bacterial cellulose by Acetobacter xylinumwith an air-lift reactor

Bacterial cellulose was produced by Acetobacter xylinum subsp. surcrofermentans BPR2001 in a 50 liter air-lift reactor using fructose as the main carbon source. When air was supplied, the production of the cellulose was only 2.3 g/l in 80 h but when O -fortified air was supplied, the cellulose concentration increased to 5.63 g/l in 28 h and the productivity of the cellulose in an air-lift reactor with O -fortified air supply was comparable to that in a mechanically agitated jar fermenter.     

AFM observation of band-like cellulose assemblies produced by Acetobacter xylinum

We succeeded in estimating the thickness of band-like cellulose assemblies by combined use of atomic force microscopy (AFM) and transmission electron microscopy. The thickness of "dense" band-like cellulose assemblies was estimated at 20-30 nm from their AFM height profiles, which was several times greater than that of "coarse" band-like and ribbonlike cellulose assemblies. On the basis of these results, the folded- chain model previously proposed was discussed, and a different organization of TC subunits was suggested for the "dense" band-like cellulose assembly.     

Bacterial cellulose production by Acetobacter xylinum in a 50-L internal-loop airlift reactor

Bacterial cellulose (BC) production was realized in a batch cultivation of Acetobacter xylinum subsp. sucrofermentans BPR2001 in a 50-L internal-loop airlift reactor. When the bacterium was cultivated with air supply, 3.8 g/L of BC was produced after 67 hours. When oxygen-enriched gas was supplied, the concentration of BC was doubled and the production rate of BC was 0.116 g/L. h, which was two times higher than that of air-supplied culture and comparable to that in a mechanically agitated stirred-tank fermentor. Bacterial cellulose produced by the airlift reactor formed a unique ellipse pellet (BC pellet), different from the fibrous form which was produced in an agitated stirred-tank fermentor. The BC-pellet suspension was demonstrated to have a higher volumetric oxygen transfer coefficient than the fibrous BC suspension in a 50-L internal-loop airlift reactor. The mixing time of BC-pellet suspension in the airlift reactor was also shorter than that in water.     

Enhancement of cellulose pellicle production by constitutively expressing vitreoscilla hemoglobin in Acetobacter xylinum

Vitreoscilla hemoglobin (VHb) gene driven by the constitutive bla promoter was expressed in the cellulose-producing Acetobacter xylinum. The expressed VHb was biochemically active and could enhance cell growth in a shaken culture containing cellulase. VHb-expressing A. xylinum (VHb+) exhibited a specific growth rate 50% higher than that of the host strain (VHb-). Probably because of its faster growth rate, the size of tentacled cellulose beads produced by VHb+ was about 20% of that produced by VHb- after 2 days cultivation in a shake-flask. When cultured statically, the amount of cellulose pellicle produced by VHb+ could be 2-fold that produced by VHb-. Cellulose pellicle concentration of 11 g/L was obtained for VHb+, whereas 6 g/L was obtained for VHb- after 6 days of microaerobic incubation.     

醋酸杆菌发酵生产细菌纤维素的研究进展

简要介绍醋酸杆菌发酵生产纤维素研究进展,内容包括：产纤维素的微生物、醋酸菌纤维素的结构特点、生物合成途径、一般提取处理及分析测定方法、商业用途、工业化发酵生产醋酸菌纤维素过程中存在的主要问题及发展前景。     

Identification of a new gene in an operon for cellulose biosynthesis in Acetobacter xylinum

DNA sequencing of the region downstream of the cellulose synthase catalytic subunit gene of Acetobacter xylinum led to the identification of an open reading frame coding for a polypeptide of 86 kDa. The deduced amino acid sequence of this polypeptide matches from position 27 to 40 with the N-terminal amino acid sequence determined for a 93 kDa polypeptide that copurifies with the cellulose synthase catalytic subunit during purification of cellulose synthase. The cellulose synthase catalytic subunit gene and the gene encoding the 93 kDa polypeptide, along with other genes probably, are organized as an operon for cellulose biosynthesis in which the first gene is the catalytic subunit gene and the second gene codes for the 93 kDa polypeptide. The function of the 93 kDa polypeptide is not clear at present, however it appears to be tightly associated with the cellulose synthase catalytic subunit. Sequence analysis of the polypeptide shows that it is a membrane protein with a signal sequence at the N-terminal end and a transmembrane helix in the C-terminal region for anchoring it into the membrane.     

Effect of various carbon and nitrogen sources on cellulose synthesis by Acetobacter xylinum

The effect of various carbon and nitrogen sources on cellulose membrane production by Acetobacter xylinum was evaluated. Among the carbon sources, sucrose, glucose and mannitol were found to be suitable for optimum levels of cellulose production. The strain was able to utilize a wide range of protein and nitrogen sources such as peptone, soybean meal, glycine, casein hydrolysate, and glutamic acid for cellulose synthesis. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of pellicle proteins (PP) revealed electrophoretic bands of molecular masses in the range of 116–20 kDa. Furthermore, the strain can be useful for the removal of various nitrogenous and carbon substrates present in waste waters.