Origin

Arthrobacter ureafaciens ※1-※2

Appearance

White amorphous powder.

Activity

More than 80 units/mg protein for N-acetylneuraminyllactose (NANA-lactose).
More than 20 units/mg protein for bovine submaxillary mucin. [at pH5.0]
More than 40 units/mg protein for colominic acid. [at pH4.5]
More than 60 units/mg protein for bovine brain ganglioside. [at pH4.0]

Unit definition

One unit is the amount of enzyme required to liberate 1 μmol of N-acetylneuraminic acid (NANA) per minute at pH5.0 at 37 ºC, using N-acetylneuraminyllactose as a substrate.

Storage

Stable for one year when stored below 5 ºC.
For prolonged storage, keep at -20 ºC.

Contaminants

Enzyme activities mentioned below cannot be detected.※1

Protease , N-Acetylneuraminic acid aldolase,
Glycosidase such as

α-Glucosidase	β-Glucosidase	α-Galactosidase
β-Galactosidase	α-Mannosidase	α-Fucosidase
N-Acetyl-α-glucosaminidase		N-Acetyl-β-glucosaminidase
N-Acetyl-α-galactosaminidase		N-Acetyl-β-galactosaminidase
N-Acetyl-α-mannosaminidase		N-Acetyl-β-mannosaminidase

Properties

Molecular weight	Approx. 52,000 Da (gel filtration, SDS-PAGE)
Optimum pH	3.8 – 4.4 (for bovine brain ganglioside)
pH stability	3.5 – 10.0
Thermal stability	below 60 ºC (pH5.0, 20 min)
Substrate specificity	sozyme S cleaves α(2→3), α(2→6) and α(2→8) linkages of N-acetylneuraminic acid in glycoconjugates. In the absence of detergents and calcium ion, the enzyme hydrolyzes N-acetylneuraminosyl moiety of polysialogangliosides and produces monosialoganglioside GM1, while in the presence of detergents (Na-cholate, Triton X-100 etc.), GM1 is further desialylated to asialoganglioside GA1. the character of the enzyme is similar to Vibrio cholerae neuraminidase except requirement of Ca2+ in the latter.

Assay method

Standard assay system is composed of 50 μl of substrate solution (4mg/ml, NANA-lactose), 50 μl of 200 mM acetate buffer (pH5.0) and 100 μl of enzyme solution. Reaction is carried out at 37 ºC for 10 minutes and NANA liberated is determined by thiobarbituric acid (TBA) method.※2-※3

References

1. ※1. Y. Ohta, Y. Tsukada and T. Sugimori, J. Biochem., 106, 1086 (1989)
2. ※2. Y. Uchida, Y. Tsukada and T. Sugimori, J. Biochem., 82, 1425 (1977)
3. ※3. L. Warren, J. Biol. Chem., 234, 1971 (1959)

Package

1 unit, 5 units