Improved Production of Mannanase byStreptomyces lividans (original) (raw)
Molekuliarnaia genetika, mikrobiologiia i virusologiia, 2012
Mannanases can be useful in the food, feed, pulp and paper industries. In this research a Bacillus subtilis strain (named Bs5) which produced high-level beta-mannanase was isolated. Maximum level of beta-mannanase (1231.41 U/ml) was reached when Bacillus subtilis Bs5 was grown on konjac powder as the carbon source for nine hours at 32 degrees C. The beta-mannanase was a typical cold-active enzyme and its optimal temperature of 35 degrees C was the lowest among those of the known mannanases from bacteria. In addition, the optimal pH was 5.0 and much wide pH range from 3.0-8.0 was also observed in the beta-mannanase. These properties make the beta-mannanase more attractive for biotechnological applications. The DNA sequence coding the beta-mannanase was cloned and the open reading frame consisted of 1089 bp encoding 362 amino acids. A phylogenetic tree of the beta-mannanase based on the similarity of amino acid sequences revealed that the beta-mannanase formed a cluster with the beta-...
SpringerPlus, 2014
The mannanase gene of B. circulans NT 6.7 was cloned and expressed in an Escherichia coli expression system. The B. circulans NT 6.7 mannanase gene consists of 1,083 nucleotides encoding a 360-amino acid residue long polypeptide, belonging to glycoside hydrolase family 26. The full-length mannanase gene including its native signal sequence was cloned into the vector pET21d and expressed in E. coli BL21 (DE3). β-Mannanase activities in the culture supernatant and crude cell extract were 37.10 and 515 U per ml, respectively, with most of the activity in the cell extract attributed to the periplasmic fraction. In contrast, expression of mannanase was much lower when using the B. circulans NT 6.7 mannanase gene without its signal sequence. The optimum temperature of recombinant β-mannanase activity was 50°C and the optimum pH was 6.0. The enzyme was very specific for β-mannan substrates with a preference for galactomannan. Hydrolysis products of locust bean gum were various mannooligosaccharides including mannohexaose, mannopentaose, mannotetraose, mannotriose and mannobiose, while mannose could not be detected. In conclusion, this expression system is efficient for the secretory production of recombinant β-mannanase from B. circulans NT 6.7, which shows good characteristics for various applications.
Isolation of mannan-utilizing bacteria and the culture conditions for mannanase production
World Journal of Microbiology & Biotechnology, 1994
A locally isolated strain, Bacillus subtilis NM-39, was selected as an active mannan-utilizing bacterium based on high saccharifying activities on coconut residue and locust bean gum galactomannan. The optimal pH and temperature ranges for activity of the crude enzyme were 5.0 to 6.0 and 50 to 60°C, respectively. The organism gave maximum mannanase activity when grown in liquid mineral salts medium containing 1% (w/v) each of coconut residue and soybean flour, as carbon and nitrogen sources, respectively, at pH 7.0 and in aerobic growth for 28 h at 37°C. High saccharifying activity on coconut mannan was also observed.
Journal of Biotechnology, 1999
A number of wild-type isolates of Sclerotium (Athelia) rolfsii and S. coffeicola were studied for their ability to produce endo-i-1,4-mannanase, endo-i-1,4-xylanase, and endo-i-1,4-glucanase activity when grown on cellulose-or glucose-based media. Whereas the presence of the inducer cellulose was strictly necessary for increased xylanase and endoglucanase production by both S. rolfsii (208 and 599 U ml − 1 , respectively) and S. coffeicola (102 and 330 U ml − 1 , respectively), elevated activities of mannanase (up to 96.6 U ml − 1 ) were formed even when employing glucose as the only carbohydrate substrate. Significant production of mannanases as well as of auxiliary mannan-degrading enzymes (i-mannosidase, i-glucosidase, h-galactosidase, acetyl esterase) was only observed, however, under derepressed conditions, i.e. after glucose had been consumed from the medium. By applying a fed-batch strategy, in which a glucose solution was continuously fed to a cultivation of S. rolfsii CBS 191.62 so that the glucose concentration in the medium never exceeded a certain low, critical value, production of mannanase could be almost doubled as compared to a batch cultivation on glucose (462 versus 240 U ml − 1 ). Mannanase preparations produced by several S. rolfsii and S. coffeicola strains under inductive and noninductive conditions (i.e. using cellulose or glucose as the substrates, respectively) were further analyzed with respect to the patterns of isoformic mannanases formed under these different growth conditions. Multiple mannanases were secreted by all isolates investigated. Certain mannanase isoenzymes were only formed by S. rolfsii in the presence of the inducer cellulose, indicating a complex and separated regulation of the synthesis of mannanase isoenzymes in this strain.
Biotechnology Progress, 2009
To be utilized in biomass conversion, including ethanol production and galactosylated oligosaccharide synthesis, namely prebiotics, the gene of extracellular endo-β-1,4-mannanase (EC 3.2.1.78) of Aspergillus fumigatus IMI 385708 (formerly known as Thermomyces lanuginosus IMI 158749) was expressed first in Aspergillus sojae and then in Pichia pastoris under the control of the glyceraldehyde triphosphate dehydrogenase (gpdA) and the alcohol oxidase (AOX1) promoters, respectively. The highest production of mannanase (352 U mL−1) in A. sojae was observed after 6 days of cultivation. In P. pastoris, the highest mannanase production was observed 10 h after induction with methanol (61 U mL−1). The fold increase in mannanase production was estimated as ∼12-fold and ∼2-fold in A. sojae and P. pastoris, respectively, when compared with A. fumigatus. Both recombinant enzymes showed molecular mass of about 60 kDa and similar specific activities (∼350 U mg−1 protein). Temperature optima were at 60°C and 45°C, and maximum activity was at pH 4.5 and 5.2 for A. sojae and P. pastoris, respectively. The enzyme from P. pastoris was more stable retaining most of the activity up to 50°C, whereas the enzyme from A. sojae rapidly lost activity above 40°C. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009
Biotechnology Progress, 2009
To be utilized in biomass conversion, including ethanol production and galactosylated oligosaccharide synthesis, namely prebiotics, the gene of extracellular endo-b-1,4-mannanase (EC 3.2.1.78) of Aspergillus fumigatus IMI 385708 (formerly known as Thermomyces lanuginosus IMI 158749) was expressed first in Aspergillus sojae and then in Pichia pastoris under the control of the glyceraldehyde triphosphate dehydrogenase (gpdA) and the alcohol oxidase (AOX1) promoters, respectively. The highest production of mannanase (352 U mL À1 ) in A. sojae was observed after 6 days of cultivation. In P. pastoris, the highest mannanase production was observed 10 h after induction with methanol (61 U mL À1 ). The fold increase in mannanase production was estimated as 12−foldand12-fold and 12−foldand2-fold in A. sojae and P. pastoris, respectively, when compared with A. fumigatus. Both recombinant enzymes showed molecular mass of about 60 kDa and similar specific activities ($350 U mg À1 protein). Temperature optima were at 60 C and 45 C, and maximum activity was at pH 4.5 and 5.2 for A. sojae and P. pastoris, respectively. The enzyme from P. pastoris was more stable retaining most of the activity up to 50 C, whereas the enzyme from A. sojae rapidly lost activity above 40 C.
Engineering in Life Sciences, 2016
Eighty-eight fungi isolated from soil and decaying organic matter were screened for mannanolytic activity. Twenty-eight fungi produced extracellular mannanase on locust bean gum as evidenced by zone of hydrolysis produced on mannan agar gel. Six prominent producers, including four Fusarium species namely Fusarium fusarioides NFCCI 3282, Fusarium solani NFCCI 3283, Fusarium equiseti NFCCI 3284, Fusarium moniliforme NFCCI 3287 with Cladosporium cladosporioides NFCCI 3285 and Acrophialophora levis NFCCI 3286 produced the β-mannanase in the range of 84-140 nkat/mL. All these grew well on particulate substrates in solid-state fermentation (SSF), producing relatively higher titers on mannan-rich palm kernel cake (PKC) and copra meal. Two high yielding strains, F. equiseti (1747 nkat/gds) and A. levis (897 nkat/gds) were selected for statistical optimization of mannanase on PKC. Interaction of two critical solid state fermentation parameters, pH and moisture on mannanase production by these two molds was studied by response surface method. Optimized production on PKC resulted in three-to fourfold enhancement in enzyme yield was observed in case of F. equiseti (5945 nkat/gds) and A. levis (4726 nkat/gds). HPLC analysis of mannan hydrolysate indicated that F. equiseti and A. levis mannanase performed efficient hydrolysis of konjac gum (up to 99%) with exclusive mannooligosaccahride (DP of 4) production. A seminative SDS-PAGE revealed that A. levis and F. solani produced three isoforms, F. moniliforme produced two isoforms while F. fusarioides, F. equiseti, and C. cladosporioides produced a single enzyme.
Enhanced β-mannanase production from alternative sources by recombinantAspergillus sojae
Acta Alimentaria, 2016
β-mannanases can degrade galactomannans to mannose and it has been used in various application areas. The aim of this study was to produce the β-mannanase from carob pod extract including different nitrogen sources. The best operation values for fermentation were determined to be 8% initial sugar concentration with 0.5% yeast extract, 100 r.p.m., and 7% inoculation rate, which yielded the maximum β-mannanase activity as 423.60 U ml-1. Effects of nitrogen sources on β-mannanase activity were also studied and it reached 695.6 U ml-1 by using 0.5% of ammonium nitrate as the nitrogen source at the determined optimum conditions. Results also showed that meat bone meal and soybean meal could be used as cost effective nitrogen sources based on achieved β-mannanase activity.
Microbial Cell Factories, 2010
Background: Mannans are one of the key polymers in hemicellulose, a major component of lignocellulose. The Mannan endo-1,4-β-mannosidase or 1,4-β-D-mannanase (EC 3.2.1.78), commonly named β-mannanase, is an enzyme that can catalyze random hydrolysis of β-1,4-mannosidic linkages in the main chain of mannans, glucomannans and galactomannans. The enzyme has found a number of applications in different industries, including food, feed, pharmaceutical, pulp/paper industries, as well as gas well stimulation and pretreatment of lignocellulosic biomass for the production of second generation biofuel. Bacillus licheniformis is a Gram-positive endospore-forming microorganism that is generally non-pathogenic and has been used extensively for large-scale industrial production of various enzymes; however, there has been no previous report on the cloning and expression of mannan endo-1,4-βmannosidase gene (manB) from B. licheniformis.