Ke Zheng

Sphingobacterium bambusae sp. nov., isolated from soil of bamboo plantation

A Gram-negative, non-motile, non-spore-forming bacterial strain designated IBFC2009T was isolated from soil of a bamboo plantation. The strain could grow at 11°C∼39°C, pH 6.0–9.0, and in the presence of 0∼5% NaCl. Based on 16S rRNA gene sequence analysis, Strain IBFC2009T belonged to the genus Sphingobacterium and showed the highest sequence similarity of 94.6% (S. composti T5-12T) with the type strains within the genus. The major fatty acids were summed feature 3 (iso-C15:0 2-OH and/or C16:1ω7c, 34.4%), iso-C15:0 (22.4%), C16:0 3-OH (15.2%), and iso-C17:0 3-OH (12.8%). The G+C content of the genomic DNA was 41.0 mol%. According to the phenotypic and genotypic characteristics, Strain IBFC2009T should represent a novel species of the genus Sphingobacterium, for which the name Sphingobacterium bambusae sp. nov. is proposed. The type strain is IBFC2009T (=CCTCC AB 209162T =KCTC 22814T).

Purification and characterization of a xylanase from Bacillus subtilis isolated from the degumming line.

Xylanases catalyze the hydrolysis of xylan, a major hemicellulose component of cell wall besides cellulose in most plant species. To extract cellulose fibers, it will be invaluable to screen for more effective xylanase‐producing microorganisms. In this paper a new strategy for easy screening of xylanase‐producing strains from the degumming line was presented. Using this strategy, a weak‐acidic, cellulase‐free xylanase from Bacillus subtilis has been isolated, purified and characterized. The xylanase showed high specific activity (36,633.4 U/mg), presented stable characteristics and can be separated and purified simply, with molecular weight 23.3 kD, pI 9.63. It reached its optimal activity at pH 5.8 and 60 °C, and retained over 80% of its maximal activity after pre‐incubation at temperature 60 °C or pH 4.6 ∼ 6.4. Also, a two‐step purification procedure based on the combination of ultrafiltration and gel filtration chromatography was introduced and described, achieving 17‐fold purification with 68.11% yield. (© 2012 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

A rapid process of ramie bio-degumming by Pectobacterium sp. CXJZU-120

Factory tests for rapid process of ramie bio-degumming by Pectobacterium sp. CXJZU-120 were conducted and evaluated comparing the processes of traditional chemical degumming and bio-chemical degumming. Results revealed that over 90% of the gum in raw ramie could be removed only with Pectobacterium sp. CXJZU-120 in 6 h. The rapid process was not only suitable for the extraction of ramie fibers from different grades of raw material and retaining the inherent morphological structures and textile properties, but also could reduce the production cost up to 20.5%, raise resource utilization by more than 50% and reduce pollution charge by more than 80% compared with the traditional chemical degumming. It is a breakthrough in the degumming of ramie and has great application potential in the extraction of herbaceous fiber materials.

Diversity and characterization of ramie-degumming strains

Ramie (Boehmeria nivea and Boehmeria tenacissima) is a widely used fiber crop. Traditional water retting or chemical boiling method performed in order to extract ramie fiber seriously pollute the environment and severely damage the fiber, so biological method is the general trend of the fiber-extracting industry. Some strains (687), involving 26 genera and 43 species, were collected from the three samples, which produce hydrolyzed circles in the selective culture medium in order to detect the degumming effect and to compare the enzyme activity. Among these strains, 13 of them did not produce cellulase and had a ramie decreasing weight rate above 25 %, which were regarded as efficient ramie-degumming strains named from R1 to R13. R1 to R13 belonged to Amycolata autotrobutylicun, Bacillus subtilis, Clostridium acetobutylicum, Bacillus subtilis, Rhizobium leguminosarum, Bacteroides finegoldii, Streptomyces lividans, Bacillus amyloliquefaciens, Clostridium acetobutylicum, Pseudomonas brassicacearum, Bacillus pumilus, Bacillus licheniformis, Pectobacterium wasabiae respectively. Bacteroides sp., Rhizobium sp. and Pseudomonas sp. were firstly reported to be used in ramie-degumming. At the same time, the pectinase was the key enzyme in the ramie-degumming process.

Purification and Characterization of a Thermostable β-Mannanase from Bacillus subtilis BE-91: Potential Application in Inflammatory Diseases

β-mannanase has shown compelling biological functions because of its regulatory roles in metabolism, inflammation, and oxidation. This study separated and purified the β-mannanase from Bacillus subtilis BE-91, which is a powerful hemicellulose-degrading bacterium using a “two-step” method comprising ultrafiltration and gel chromatography. The purified β-mannanase (about 28.2 kDa) showed high specific activity (79, 859.2 IU/mg). The optimum temperature and pH were 65°C and 6.0, respectively. Moreover, the enzyme was highly stable at temperatures up to 70°C and pH 4.5–7.0. The β-mannanase activity was significantly enhanced in the presence of Mn2+, Cu2+, Zn2+, Ca2+, Mg2+, and Al3+ and strongly inhibited by Ba2+ and Pb2+. K m and V max values for locust bean gum were 7.14 mg/mL and 107.5 μmol/min/mL versus 1.749 mg/mL and 33.45 µmol/min/mL for Konjac glucomannan, respectively. Therefore, β-mannanase purified by this work shows stability at high temperatures and in weakly acidic or neutral environments. Based on such data, the β-mannanase will have potential applications as a dietary supplement in treatment of inflammatory processes.

An alkaline pectate lyase D from Dickeya dadantii DCE-01: clone, expression, characterization, and potential application in ramie bio-degumming

Pectinase plays a crucial role in ramie bio-degumming. A pectate lyase gene (pel4J4) from the high-efficiency degumming bacteria Dickeya dadantii DCE-01 of bast fibers was cloned and connected to pET28a, and then the recombinant plasmid was successfully transformed into Escherichia coli BL21(DE3). The pectate lyase (Pel4J4) induced was purified by ultrafiltration and Sephadex G-100 gel chromatography. The enzymatic properties of Pel4J4 were studied in detail. pel4J4 (GenBank accession number: KC900167) had a sequence length of 1179 bp, encoding 392 amino acids. The extracellular pectate lyase activity of pET28a-pel-BL was up to 204.4 IU/mL. The optimal temperature and pH of the purified Pel4J4 were 55℃ and 8.5, respectively. The stable temperature and pH of Pel4J4 activity were 45℃ and 8.5–10.0, respectively. The catalytic activity is Ca2+ dependent and promoted by 1 mmol/L Zn2+, Fe3+, Ca2+, and NH4+, but seriously inhibited by Cu2+ and Pb2+. The optimal substrate is citrus pectin with more than 85% esterification. The heat-resistant alkaline Pel4J4 could strongly degrade natural ramie pectin, indicating a promising application prospect in ramie bio-degumming.

Bio-degumming technology of Apocynum venetum bast by Pectobacterium sp. DCE-01

Apocynum venetum fiber is known as the “king of wild fibers”. The traditional chemical degumming method for A. venetum limits the development of the industry due to serious pollution and fiber damage. By machine rolling preprocessing, bacteria culture, soaking fermentation, inactivation, water scrubbing, and drying under fermentation conditions of bath ratio 1:15, inoculum size 2%, temperature 33℃, and fermentation time 16 h, A. venetum fiber could be extracted from A. venetum bast. Strain Pectobacterium sp. DCE-01 simultaneously secreted pectinase, mannanase, and xylanase, which can decompose the main components of A. venetum non-cellulose. The catalytic activities of pectase, mannanase, xylanase, and cellulase at 16 h were 12.47, 7.56, 6.02, and 2.82 U/ml respectively. Compared with chemical degumming, bio-degumming exhibited 22.56% higher residual gum content, 18.14% higher breaking strength, and 44.33% lower chemical oxygen demand.

Diversity and Characteristics of Kenaf Bast Degumming Microbial Resources

ABSTRACT Kenaf is one of the most important natural fiber crops. Traditional degumming with water retting causes serious environmental pollution and reduces the quality of fiber products. The development of kenaf industry is hindered by high production cost. Microbial degumming is suitable for kenaf bast degumming because of its high efficiency, energy conservation, low pollution, and high quality, among others. Through enrichment and screening from water sample, soil sample, and humus sample, we concentrated and identified 92 bacterial strains that could degrade kenaf bast colloid. The strains belonged to 11 genera and 16 species. Five of these strains did not produce cellulase and the weight loss rate of the kenaf bast fiber raw material was more than 20%. These strains belonged to Bacillus subtilis, Paenibacillus polymyxa, Clostridium acetobutylicum, Bacillus alcalophilus, and Erwinia chrysanthemi and were assigned with serial numbers from K1–K5, respectively. This study is the first to report the function of Bacillus pumilus, B. alcalophilus, Clostridium tertium, Brevibacillus brevis, Pectobacterium carotovora, E. chrysanthemi, and Tyromyces subcaesius in kenaf bast degumming. Pectinase and mannanase were the key enzymes in the degumming of kenaf bast.

A novel endo-β-1,4-xylanase GH30 from Dickeya dadantii DCE-01: Clone, expression, characterization, and ramie biological degumming function:

Xylanase plays an important role in the hydrolysis of hemicellulose and has gained much attention in the field of biological degumming. The research for xylanases with cellulase-free and high activity for biological degumming has intensified in recent years. In the present research, heterologous expression of a novel endo-β-1,4-xylanase (GH30) from Dickeya dadantii DCE-01 in Escherichia coli BL21 (DE3) was reported. Biochemical characterization of the enzyme and a potential application in ramie biological degumming was discussed. The results showed that the xylanase gene consists of 1251 nucleotides, belonging to glycoside hydrolase family 30 (GH30). The optimal activity of the xylanase was observed at 50℃ and a pH value of 6.4. The Km and Vmax values for beechwood xylan were 14.25 mg/mL and 296.6 μmol/mg, respectively. The catalytic activity was enhanced by addition of 1 mM Cu2+, Ca2+, Mg2+, and K+. The recombinant enzyme was specific for xylan substrates. The enzyme exhibited hydrolytic activity toward ramie hemicellulose. The recombinant xylanase could be effectively applied to ramie degumming.