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CitST is a typical two-component regulatory system that can regulate citrate utilization and is involved in the regulation of carbohydrate metabolism in Bacillus subtilis. B. subtilis is a kind of dominant species to bio-degum ramie fibers, which could degrade and utilize non-cellulose components as a carbon source during bio-degumming. This study aimed to explore the regulatory function of response regulator CitT from CitST two-component system when B. subtilis was used for extracting ramie fibers. The results showed that gene knocked-out of citT could significantly decrease the bio-degumming ability of B. subtilis. The enzymatic activity mensuration of functional enzymes revealed that deficient in CitT would remarkably reduce the enzymatic activity of pectinase, but there was a little difference in transcriptional levels of their encoding genes with qRT-PCR analysis, which further demonstrated that it was a lack of correlation between mRNA expression and enzymatic activities. The data also reflected that CitT specifically regulated the pectin degradation in bio-degumming of ramie fibers by analyzing the polysaccharide degradation rates of gum components. In conclusion, it was shown for the first time that CitT probably positively regulated the bio-degumming of ramie fibers and was an important regulator in pectin degradation. [ABSTRACT FROM AUTHOR]

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In order to obtain a thermostable pectate lyase for ramie degumming, a rational design based on structural analysis was carried out on a novel pectate lyase (Pel419) derived from the Dickeya Dadantii DCE-01 for high-efficiency ramie degumming. A total of five potential amino acid sites were chosen to replace residues. Then, the mutant enzymes were subjected to the heterologous expressions in Escherichia coli and their enzymatic characteristics were determined. The optimal reaction temperature for the five mutants kept consistent with that for the wild type. The enzyme activity and thermal stability of mutant V52A were significantly improved. Meanwhile, the weight loss rate obtained by V52A with the best enzymatic characteristics in the ramie degumming process at 50 °C is comparable with that obtained by commercial cotton-ramie processing pectinases, indicating that V52A was a potential industrial enzyme that could be applied to large-scale ramie degumming. In this study, the biological functions of conservative residues of Pel419 were preliminarily explored. The mutant V52A with both enzymatic activity and improved heat resistance was acquired, providing a superior material for developing enzyme preparations of ramie degumming, and rendering an effective method for the rational design aiming to improve the thermostability of pectate lyase. [ABSTRACT FROM AUTHOR]

Microbial treatment of natural fibers is a preferred alternatives in ramie degumming process as compared to conventional chemical degumming technology with alkaline treatment at high temperature. Usually, it is difficult to degrade the noncellulosic components of ramie bast completely with a single strain in the mono-culturing degumming system. Microbial coculture strategy has been widely used in biotechnology, especially in the degradation of biological materials. In the present study, five strains for ramie degumming were identified as Dickeya dadantii strain DCE-01, Bacillus subtilis strain 1101, Bacillus sp. B6, Bacillus cereus strain B7, and Bacillus cereus strain B8 by sequencing of 16 S rRNA genes. Enzymatic activities characterization, ramie degumming experiments with mono-culturing systems, and the evaluation of strain compatibility showed that group DCE-01/B7 were the most suitable pairs for the construction of bacterial coculturing system. After preliminary optimization of coculturing fermentation condition, the weight loss weight ratio of ramie fibers was 27%, which was increased 13.6% and 8.9% while compared to mono-culturing systems of strain B7 and DCE-01 respectively. These data highlighted the benefit of microbial coculture strategy in the application of ramie degumming process, and indicated a higher-efficiency alternative for the microbial degumming in the textile industry. [ABSTRACT FROM AUTHOR]

To explore high-quality microbial resources with the capability of ramie degumming, we collected soil samples from rotten ramie and straw heaps. After enrichment culture by ramie raw materials, bacterial strains with the potential ramie-degumming function were screened using a pectin-hydrolysis plate. Dominant bacteria were identified by combining colonial morphological characteristics with the molecular biology method, and their ramie-degumming effects were verified through comprehensive biological degumming indices. Results demonstrated that Bacillus aryabhattai, Bacillus thuringiensis, Lysinibacillus fusiformis, and Acidovorax temperans were successfully obtained. The highest pectinase activity, 98.2 U/mg, was found by A. temperans. B. thuringiensis showed the best ramie-degumming effect. The residual gum content, single-fiber linear density, and bundle-breaking strength of the degummed ramie fiber treated with B. thuringiensis were 8.32%, 6.80 dtex, and 7.84 cN/dtex, respectively. The residual gum content of the ramie fiber treated with B. thuringiensis met the textile requirement (<10%), and the values of all other indicators were also satisfactory. Therefore, B. thuringiensis was an excellent strain for ramie degumming, indicating potential industrial applications. [ABSTRACT FROM AUTHOR]

Ramie (Boehmeria nivea L.), a perennial herb, is an important bast fiber plant. Its fiber with the advantages of attractive luster, high tenacity, enhanced strength, and good microbial resistivity is well known as the queen of natural fibers. The abundant cellulose fibers in ramie raw materials are stuck tightly by gums consisting of pectic substances, hemicelluloses, and little lignin. The gum should remove from the ramie raw material through degumming process to separate fibers, unveil unique fiber properties, and improve fiber-spinning ability to fulfill textile requirements. Low degumming efficiency and high environmental pollution are the major problems hindering the utilization of ramie fibers. Ramie degumming involves the degradation of pectin and hemicelluloses, which requires chemical, physical, biological treatment, or a combination of several treatments. No stereotyped parameters of the given degumming method have been yet established for the extraction of textile-grade ramie fibers. This review evaluated integrated methodology involving chemical, physical, biological and biochemical methods to degum raw ramie and obtain textile-grade refined fibers. [ABSTRACT FROM AUTHOR]

Pectinase plays a crucial role in ramie bio-degumming. A pectate lyase gene (pel 4J4) 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. pel 4J4 (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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

To improve the thermal stability of pectate lyase for ramie degumming, we modified the novel pectate lyase gene (pelG403) derived from the Dickeya dadantii DCE-01 high-efficiency ramie degumming strain by site-directed mutagenesis. Twelve mutants were acquired, wherein a prospective mutant (A129V) showed better enzyme activity and thermal stability. Compared with the wild type (PelG403), the specific enzyme activity and the optimal reaction temperature of A129V in the fermentation broth increased by 20.1%, and 5 °C, respectively. Under the conditions of 55 °C and pH 9.0, the weightlessness rate of ramie raw materials of A129V increased by 6.26%. Therefore, this study successfully improved the enzyme activity and heat resistance of PelG403 in an alkaline environment, which may contribute to the development of enzyme preparations and the elucidation of the mechanism for ramie bio-degumming. [ABSTRACT FROM AUTHOR]