CsAra51A

Ex-Ara0142

(EC.3.2.1.55)exo-α-Arabinofuranosidase

CAZy Family: GH51

PROPERTIES

1.ELECTROPHORETIC PURITY

-Single band on SDS-gel electrophoresis (MW ~58 kDa)

Figure 1. Electrophoresis analysis ofCsAra51A. 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, CsAra51A purified from Ni sepharose fastflow column.

2.SPECIFIC ACTIVITY

28.2 U/mg protein (on pNP-α-Araf) at pH 5.5 and 80°C.

One Unit of pNP-α-Arafactivity is defined as the amount of enzyme required to release 1 μmol of p-nitrophenyl per minute from pNP-α-Araf (2 mM) in citric acid buffer (50 mM, pH 5.5)

3.RELATIVE RATES OF HYDROLYSIS OF SUBSTRATES

Table 1. Relative activity of CsAra51A on different substrates^a.

^aReactions were performed with 5 mM substrate, pH5.5, at 80°C for 5 min.

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

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

Figure 2. Effect of enzyme concentration on ginsenoside Rd production from ginsenoside Rc byCsAra51A. (A) Effect of enzyme concentration on ginsenoside Rd production. (B) Ginsenoside Rd (●) production from ginsenoside Rc (○) under the optimum conditions. Data represent the means of three experiments and error bars represent standard deviation

4.PHYSICOCHEMICAL PROPERTIES

pH Optima: 5.5

Temperature Optima: 80°C

Figure2.Effects of pH and temperature on the activity of CsAra51A. (a) pH effect:. (b) Temperature effect.

Figure 3. Thermal inactivation of CsAra51Aat temperatures of 70°C (□), 75°C (■), 80°C (○), and 85°C (●) for varying time periods.

5.STORAGE CONDITIONS

The enzyme should be stored at -20°C. For assay, this enzyme should be diluted in sodiumacetate (20 mM) pH5.5. Swirl to mix the enzyme immediately prior to use.

6. REFERENCES

[1] Lim Y R, Yoon R Y, Seo E S, et al. Hydrolytic properties of a thermostable α‐l‐arabinofuranosidase from Caldicellulosiruptor saccharolyticus[J]. Journal of applied microbiology, 2010, 109(4): 1188-1197.

[2]Shin K C, Lee G W, Oh D K. Production of Ginsenoside Rd from Ginsenoside Rc by ${\alpha}-{\small {L}} $-Arabinofuranosidase from Caldicellulosiruptor saccharolyticus[J]. Journal of microbiology and biotechnology, 2013, 23(4): 483-488.