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Table 1 Purification of the recombinant Tth xynB3 β-xylosidase

From: Biochemical properties of a novel thermostable and highly xylose-tolerant β-xylosidase/α-arabinosidase from Thermotoga thermarum

Purification step Total volume (mL) Total activity (U) Total protein (mg) Specific activity (U/mg) Recovery (%) Purification (fold)
Crude extracta 10 2394 122 20 100 1
Heat treatmentb 10 2155 39 55 90 2.8
Ni affinity chromatographyc 1 2011 17 116 84 5.8
  1. a The recombinant strain was grown in LB medium (200 ml) with 1 μg ampicillin/ml at 37°C to OD600 0.8 and was incubated further with isopropyl-β-thiogalactopyranoside (IPTG) for 12 h. The cells were harvested by centrifugation at 10,000 g for 15 min at 4°C and resuspended in 10 ml imidazole buffer (10 mL of 5 mM imidazole, 0.5 mM NaCl, and 20 mM Tris–HCl buffer, pH 7.9), followed by sonication.
  2. b The cell extracts after sonication were heat treated at 70°C for 30 min, and then cooled in an ice bath, centrifuged at 15,000 g for 20 min at 4°C and the supernatant was kept.
  3. c The obtained supernatants were loaded on to an immobilized metal affinity column (Novagen, USA), and eluted with 0.4 M imidazole, 0.5 M NaCl, and 20 mM Tris–HCl buffer (pH 7.9).
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