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Table 2 Specific activity of the purified β-glucosidases towards various cello-oligosaccharides (degree of polymerisation [DP] from 2 to 5) and carbohydrate polymers

From: Microbial β-glucosidases from cow rumen metagenome enhance the saccharification of lignocellulose in combination with commercial cellulase cocktail

Substrate

Specific activity (U mg-1)

Pure β-glucosidases

SRF2g14

SRF2g18

LAB20g4

LAB25g2

Cellobiosea

369.9 ± 0,04

852.8 ± 0.03

305.9 ± 0.05

1537. 2 ± 0.01

Cellotriosea

317.6 ± 0.04

789.4 ± 0.03

60.2 ± 0.01

2170. 5 ± 0.02

Cellotetraosea

343.0 ± 0.01

944.4 ± 0.01

283.9 ± 0.01

2154. 3 ± 0.01

Cellopentaosea

397.4 ± 0.001

1154.8 ± 0.03

313.5 ± 0.02

2682. 6 ± 0.01

Lichenana

60. 8 ± 0.13

n.d

22.8 ± 0.16

n.d

 

E. coli BL21 (DE3) cell extracts expressing β-glucosidasesb

p NPβGluc

0.39

5.1×10-3

0.40

1.5

Cellobiosed

115.2

16.2

105.9

231.5

  1. aReaction assays contain 2 μg pure enzyme, 200 μL 50 mM sodium acetate buffer (pH 5.6) and 100 μL of a 100 mM cellobiose stock solution (final concentration of cellobiose of 20 mM). For lichenan a concentration of 10 mg ml-1 was used. Samples were incubated at 40°C during 5 h, after which the glucose concentration was measured using the glucose oxidase kit (Sigma Chemical Co., St. Louis, MO, USA). All of the measurements were analysed in triplicate; n.d. no activity detected.
  2. bQuantification of proteins in E. coli BL21 (DE3) cell extracts expressing β-glucosidases was performed according to BCA Protein Assay kit (Thermoscientific), and are as follows: 552, 426, 458 and 508 μg ml-1 for cell extracts of SRF2g14, SRF2g18, LAB20g4 and LAB25g2, respectively.
  3. cActivity measured by using 100 μl cell extract, 50 μl p NPβG solution (0.22% w/v) and 150 μL buffer McIlvane (100 mM, pH 5). Incubation: 20 min at 40°C, after which 250 μl Na2CO3 (1 M) was added and absorbance measured at 405 nm. Specific activity was measured as follows: mU ml-1 enzyme = A/KtV (A = absorbance at 405 nm; K = 0.0328 (value determined by calibration); V = volume in ml. The standard deviation was lower than 5%.
  4. dReaction assays contain 200 μl enzyme extract, 200 μl McIlvaine buffer and 100 μl of a 100 mM cellobiose stock solution (final concentration of cellobiose of 20 mM). Sample were incubated at 40°C during 4 h, alter which the glucose concentration was measured using the glucose oxidase kit (Sigma Chemical Co., St. Louis, MO, USA). The standard deviation was lower than 5%.
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Biotechnology for Biofuels and Bioproducts

ISSN: 2731-3654

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