Products

Zymolyase-20T / Zymolyase-100T

Zymolyase

Zymolyase®, produced by a submerged culture of Arthrobacter luteus(1), has strong lytic activity against living yeast cell walls(2),(3) to produce protoplast or spheroplast of various strains of yeast cells. An essential enzyme for the lytic activity of Zymolyase® is β-1,3-glucan laminaripentaohydrolase. It hydrolyzes linear glucose polymers with β-1,3-linkages and releases specifically laminaripentaose as the main and minimum product unit(4), (5), (10), (11).

Product Information

There are two preparations of Zymolyase®, Zymolyase®-20T and Zymolyase®-100T, having lytic activity of 20,000 units/g and 100,000 units/g respectively. Zymolyase®-20T is ammonium sulfate precipitate while Zymolyase®-100T is a further purified preparation by affinity chromatography(9). Lytic activity varies depending on yeast strain, growth stage of yeast, or cultural conditions(6-8). Further informations related to Zymolyase® can be obtained in the reference section below(12-16).

Product Information

Product Name Zymolyase®-20T Zymolyase®-100T
Form Lyophilized Powder
Purification Ammonium Sulfate Precipitation Affinity Chromatography
Activity 20,000 units/g 100,000 unites/g
Essential enzyme β-1,3-glucan laminaripentaohydrolase
Other activities
contained (※1)		 β-1,3-glucanase approx. 1.5 x 106 units/g approx. 1.0 x 107 units/g
protease approx. 1.0 x 104 units/g approx. 1.7 x 104 units/g
mannanase approx. 1.0 x 106 units/g approx. 6.0 x 104 units/g
Contaminants Amylase, Xylanase,
Phosphatase		 Trace amount Non detectable
Optimum pH and Temp. pH7.5, 35°C (for lysis of viable yeast cells)
pH6.5, 45°C (for hydrolysis of yeast glucan)
Stability 2°C No loss of activity was found after storage for 1 year
Heat stability 30°C 70% of the lytic activity is lost after storage for 3 months
90% of the lytic activity is lost after storage for 3 months
60°C Lytic activity is lost on incubation for 5 minutes
Specificity (Lytic Spectrum) Ashbya, Candida, Debaryomyces, Eremothecium, Endomyces, Hansenula, Hanseniaspora, Kloeckera, kluyveromyces, Lipomyces, Metschnikowia, Pichia, Pullularia, Torulopsis, Saccharomyces, Saccharomycopsis, Saccharomycodes, Schwanniomyces, etc.

(※1) See reference, Kitamura, K., Kaneko, T., Yamamoto, Y., J. Gen. Appl. Microbiol., 18, 57 (1972) as to the definition of each enzyme units.

Unit Definition

One unit of lytic activity is defined as that amount which indicates 30% of decrease in absorbance at 800 nm(A800) of the reaction mixture under the following condition.

Reaction Mixture

Enzyme solution
 1 ml (0.05-0.1 mg/ml for Zymolyase®-20T)
1 ml (0.012-0.024 mg/ml for Zymolyase®-100T)
Brewer's yeast cell suspension 3 ml (2 mg/ml)
1/15M Phosphate buffer 5 ml (pH7.5)
Distilled water 1 ml

After incubation for 2 hours at 25°C with gentle shaking, A800 of the mixture is determined. When 60% of A800 decrease, equivalent to 2 units, is observed in the reaction system, the brewer’s yeast cells are completely lysed, namely 1 unit of Zymolyase® lyses 3 mg dry weight of brewer’s yeast.

Applications

Electron microscopical photo of yeast cell

Electron microscopical photo of yeast cell

Electron microscopical photo of yeast cell
(Candida tropicals)
CW: Cell Wall
CM: Cell Membrane
M: Mitochondria
Mb: Microbody
N: Nucleus
V: Vacuole

               ↓

Electron microscopical photo of yeast cell

Yeast cell(Candida tropicalis)
after Zymolyase® treatment.

(Data courtesy of Professor Masako Osumi, Nippon Women's University)

Protocols


Preparation of 1/15 M Phosphate Buffer (1L of pH 7.5 buffer)

  1. Dissolve 19.1 g of di-sodium hydrogenphosphate 12-hydrate in distilled water to make an 800 mL solution.
  2. Dissolve 1.814 g of potassium dihydrogenphosphate in distilled water to make a 200 mL solution.
  3. Mix the two solutions.

The below phosphate buffers (pH 7.5) have been tested for compatibility.

Buffer

Concentration

Yeast Lytic activity (unit/g)

Potassium/Sodium Phosphate Buffer

1/15 M

21,000

Potassium Phosphate Buffer

5 mM

20,300

10 mM

21,400

50 mM

21,000

Sodium Phosphate Buffer

5 mM

20,500

10 mM

21,200

50 mM

21,200

References

  1. K Takagi et al.: Chemical and Pharmaceutical Bulletin?64.5 (2016): 432-438
  2. S Ikushiro et al.: Molecular pharmaceutics(2016)
  3. M Schuler et al.: Journal of Biological Chemistry(2016): jbc-M116
  4. E Tanaka et al.: Plant Pathology (2016)
  5. S Yamamoto et al: Biochemical and Biophysical Research Communications (2016)
  6. Y Ootsubo et al.: Bioscience, biotechnology, and biochemistry80.4 (2016): 748-760
  7. Y Morimoto et al.: Molecular microbiology 95.4 (2015): 706-722
  8. M Murai et al.: Biochemistry 53.24 (2014)
  9. M Asaoka et al.: Journal of biochemistry156.6 (2014)
  10. Herrero Enrique, Sanz Pascual. Sentandreu Rafael: J. Gen. Microbiol., 133 (10), 2895 (1987)
  11. Iijima Y., Yanagi S. O.: Agric. Biol. Chem., 50 (7), 1855 (1986)
  12. Shibata Nobuyuki, Kobayashi Hidemitsu, tojo Menehiro, Suzuki Shigeo: Arch. Biochem. Biophys., 251(2), 697 (1986)
  13. Iizuka Masaru, Torii Yasuhiko, Yamamoto Takehiko: Agric. Biol. Chem., 47 (12), 2267 (1983)
  14. Calza R. E., Schroeder A. L.: J. Gen. Microbiol., 129, 413 (1983)
  15. Kitamura, K.: Agric. Biol. Chem., 46, 2093 (1982)
  16. Kitamura, K.: Agric. Biol. Chem., 46, 963 (1982)
  17. Katamura, K.: J. Ferment. Technol., 60, 257 (1982)
  18. Katamura, K. and Tanabe, K.: Agric, Biol. Chem., 46, 553 (1982)
  19. Kitamura, K. and Yamamoto,.: Agric. Biol. Chem., 45, 1761 (1981)
  20. Kitamura, K., Kaneko, T. and Yamamoto, Y.: J. Gen Appl. Microbiol., 20, 323 (1974)
  21. Kaneko, T., Kitamura, K. and Yamamoto, Y.: Agric. Biol. Chem., 37, 2295 (1973)

Downloads

Zymolyase®-20T and Zymolyase®-100Tpdf (PDF 637 KB)

Ordering Information

Product Name Storage Product No. PKG Size Price
Zymolyase®-20T R 07663-91 1 g e-Nacalai.jpg
07663-04 10 g
Zymolyase®-100T R 07665-55 500 mg e-Nacalai.jpg
07665-84 5 g

Zymolyase® is a registered trademark of Kirin Holdings Company, Limited

(Storage) R: Refrigerator