Isolation and partial characterization of cellulose-degrading strain of Streptomyces sp. LX from soil

Abstract

A previous report dealt with the cloning in Escherichia coli and sequencing of both the cDNA and genomic DNA encoding a highly active xylanase (XynA) of Aureobasidium pullulans (X.-L. Li and L. G. Ljungdahl, Appl. Environ. Microbiol. 60:3160-3166, 1994). Now we show that the gene was expressed in Saccharomyces cerevisiae under the GAL1 promoter in pYES2 and that its product was secreted into the culture medium. S. cerevisiae clone pCE4 with the whole open reading frame of xynA, including the part coding for the signal peptide, had xylanase activity levels of 6.7 U ml-1 in the cell-associated fraction and 26.2 U ml-1 in the culture medium 4 h after galactose induction. Two protein bands with sizes of 25 and 27 kDa and N-terminal amino acid sequences identical to that of APX-II accounted for 82% of the total proteins in the culture medium of pCE4. These proteins were recognized by anti-APX-II antibody. The results suggest that the XynA signal peptide supported the posttranslational processing of xynA product and the efficient secretion of the active xylanase from S. cerevisiae. Clones pCE3 and pGE3 with inserts of cDNA and genomic DNA, respectively, containing only the mature enzyme region attached by a Met codon had low levels of xylanase activity in the cell-associated fractions (1.6 U ml-1) but no activity in the culture media. No xylanase activity was detected in clone pGE4, which was the same as pCE4, except that pGE4 had a 59-bp intron in the signal peptide region. A comparison of the A. pullulans and S. cerevisiae signal peptides demonstrated that the XynA signal peptide was at least three times more efficient than those of S. cerevisiae invertase or mating alpha-factor pheromone in secreting the heterologous xylanase from S. cerevisiae cells.

Predominance of Trichoderma and Penicillium in cellulolytic aerobic filamentous fungi from subtropical and tropical forests in China, and their use in finding highly efficient β-glucosidase

“…Whereas the Avicelase activity produced by an actinomycete strain isolated by Li et al was only 0.008 U/ml [31], Douglas et al isolated a fungal strain with higher Avicelase activity (0.12 to 0.17 U/ml) [32]. In the present study, 17% of the strains isolated using Whatman No.…”

Biodegradation of xanthan by newly isolated Cellulomonas sp. LX, releasing elicitor-active xantho-oligosaccharides-induced phytoalexin synthesis in soybean cotyledons

“…at 650 nm. Viscosity was measured at 40 8C with Rotational Viscometer NDJ-1 (Shanghai Balances Instrument Factory, China) as described previously [23]. Reducing sugars, calculated as glucose, were determined by the method of dinitrosalicylic acid [22].…”

“…A mixture containing equal volumes of the culture supernatant and 0.6% (w/v) of xanthan in 30 mmol/l phosphate buffer (pH 6.0) was incubated at 40 8C for 30 min and the viscosity was measured by the method described previously [23]. Appropriate controls for the viscosity decrease without addition of the culture supernatant were included.…”

“…The reaction mixture was removed at interval time points and the reducing sugar was determined by the method of dinitrosalicylic acid [22] over 120 min incubation. Simultaneously, the viscosity of the reaction mixture was measured according to the method previously described [23].…”

Production and characterization of endoglucanase secreted by Streptomyces capoamus isolated from Caatinga

“…Many studies have reported that the maximum activity of cellulases produced by bacteria has its peak in temperature ranging between 50 and 55°C, and it is interesting that in all the works cited, an activity above 55°C falls below 40% of residual activity (Ishaque and Kluepfel, 1980;Li and Gao, 1996;George et al, 2001;Saha et al, 2006;Prasad and Sethi, 2013). Even in studies with actinomycetes of the genus Streptomyces (S. halstedii, S. thermodiastaticus and S. reticuli) production peaks have shown a great cellulase activity at a temperature of 55°C (Crawford and Mccoy, 1972;Garda et al, 1997;Ramirez and Coha, 2003).…”