BoXyl31A

Ex-Xyl00161

(EC.3.2.1) exo-α-Xylosidase

CAZy Family: GH31

PROPERTIES

1.ELECTROPHORETIC PURITY

-Single band on SDS-gel electrophoresis (MW ~88 kDa)

Figure 1. Electrophoresis analysis of BoXyl31A. M, molecular weight marker (PageRuler Prestained Protein Ladder, Thermo Scientific); lane 1, culture lysate before IPTG induction; lane 2, culture lysate after IPTG induction; lane 3, BoXyl31A purified from Ni sepharose fastflow column.

2.SPECIFIC ACTIVITY

2.91 U/mg protein (on pNP-α-Xyl) at pH 7.0and 37^oC.

One Unit ofpNP-α-Xyl activity is defined as the amount of enzyme required to release one μmole ofp-nitrophenyl per minute frompNP-α-Xyl (5 mM) in phosphate buffer (20 mM) pH 7.0.

3.RELATIVE RATES OF HYDROLYSIS OF SUBSTRATES

Table 1. Relative activity ofBoXyl31A on different substrates^a.

^aReactions were performed with 5mM substrate, pH 7.0, at 37°C for 5 min.

^bAbsorption caused by released p-nitrophenol was measured at 405 nm. The relative activity on pNPαXyl was taken as 100%.

^cThe data are reported as means±standard errors from the mean for three independent experiments.

4.PHYSICOCHEMICAL PROPERTIES

pH Optima : 6.0

pH Stability: 6.0-10.0

Temperature Optima: 40°C

Temperature Stability:<45°C

Figure2. Effect of temperature and pH on the activity and stability of recombinant BoXyl31A. (A) Optimum pH. Assays were carried out at 37°C for 5 min in buffers ranging in pH from 2.0 to 11.0. The activity at optimum pH was defifined as 100%. (B) pH stability. Assays were carried out after the incubation of the enzyme in buffers ranging from pH 2.0-11.0 at 25°C.(C) Optimum temperature. Activity was measured between 20°C and 90°C at pH 7.0 for 5 min. The activity at optimum temperature was defifined as 100%. (D)Thermostability. Assays were carried out in 20 mM phosphate buffer (pH 7.0) at 37 °C for 5 min after the incubation of the enzyme a t35-55°C. The activity without heat treatment was defifined as 100%.

5.STORAGE CONDITIONS

The enzyme should be stored at -20°C. For assay, this enzyme should be diluted in phosphate buffer (20 mM) pH 7.0. Swirl to mix the enzyme immediately prior to use.