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Zymolyase®
(Zymolyase® 20T & Zymolyase® 100T)
 Zymolyase® vs Lyticase & Glusulase
Product Overview
Overview
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 beta.-1,3-glucan laminaripentaohydrolase. It hydrolyzes linear glucose polymers at beta.-1,3-linkages and releases specifically laminaripentaose as the main and
minimum product unit(4), (5), (10), (11).
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 information related to Zymolyase® can be obtained in the reference section
below(12-16).
Applications
- Protoplast/spheroplast preparation
- Yeast cell fusion
- Transformation of yeast cells
- Yeast genetics
Latest Publications
Chatterjee A (2011) Saccharomyces cerevisiae THI4p is a suicide thiamine thiazole synthase. Nature. Oct 26;478(7370):542-6.
Park JN (2011) Functional analysis of a Hansenula polymorpha MNN2-2 homologue encoding a putative UDP-N-acetylglucosamine transporter localized
in the endoplasmic reticulum. The Journal of Microbiology. Vol. 49, No. 6, pp. 1012-1017
Stringer DK and Piper RC (2011) A single ubiquitin is sufficient for cargo protein entry into MVBs in the absence of ESCRT ubiquitination.
The Journal of Cell Biology, vol. 192 no. 2 229-242
Oh J & Nislow C (2011) Signature-tagged Mutagenesis to Characterize Genes Through Competitive Selection of Bar-coded Genome Libraries.
Methods in Molecular Biology, 2011, Volume 765, Part 2, 225-252
Walker SC et. al (2011) The Dual Use of RNA Aptamer Sequences for Affinity Purification and Localization Studies of RNAs
and RNA–Protein Complexes. Methods in Molecular Biology, Volume 714, Part 5, 423-444
Product Specifications
| Form: |
Lyophilized powder |
| Purification: |
Zymolyase® -20T:(NH4)2SO4 precipitation
Zymolyase® -100T:Affinity Chromatography |
| Activity: |
Zymolyase® -20T:20,000 units/gram
Zymolyase® -100T:100,000 units/gram |
| Essential enzyme: |
beta.-1, 3-glucan laminaripentaohydrolase |
| Other activities contained: |
|
Zymolyase® - 20T |
Zymolyase® - 100T |
| B-1, 3-glucanase: |
ca. 1.5 x 106 units/g |
ca. 1.0 x 107 units/g |
| protease: |
ca. 1.0 x 104 units/g |
ca. 1.7 x 104 units/g |
| mannanase: |
ca. 1.0 x 106 units/g |
ca. 6.0 x 104 units/gf |
|
| Contaminants: |
Trace amounts of amylase, xylanase, phosphatase.
No DNase, RNase detected |
| Optimum pH & temperature: |
pH 7.5, 35 deg. C (for lysis of viable yeast cells)
pH 6.5, 45 deg. C (for hydrolysis of yeast glucan) |
| Stable pH: |
5~10 |
| Heat stability: |
The lytic activity is lost on incubation at 60 deg. C for 5 minutes. |
| Specificity (lytic spectrum)(5): |
Ashbya, Candida, Debaryomyces, Eremothecium, Endomyces, Hansenula, Hanseniaspora, Kloeckera, Kluyveromyces, Lipomyces, Metschikowia, Pichia, Pullularia, Torulopsis, Saccharomyces, Saccharomycopsis, Saccharomycodes, Schwanniomyces, etc. |
| Activators: |
SH compound such as cystein, 2-mercaptoethanol of dithiothreitol |
| Stability: |
No loss of activity was found after storage for 1 year at 4 deg. C |
Properties of Zymolyase®
Lytic Spectrum of Zymolyase®
1) Susceptible strains in low concentration (0.2 units/ml)
Ashbya, Endomyces, Kloeckera, Kluyveromyces, Pullularia, Saccharomyces
2) Susceptible strains in high concentration (2.0 units/ml)
Candida, Debaryomyces, Eremothecium, Hansenula, Hanseniaspora, Lipomyces, Metschikowia, Saccharomycopsis, Saccharomycodes, Schizosaccahromyces, Selenozyma, Trigonopsis,Wickerhamia
3) Susceptibility depending on strains
Bretanomyces, Cryptococcus, Nadsonia, Pichia, Rodosporidium, Schwanniomyces, Stephnoascus, Torulopsis
4) No susceptible strains
Bullera, Pityrosporum, Rhosotorula, Sporidiobolus, Sporobolomyces, Stetigmatomyces, Trichosporon
Assay for Enzyme Activity
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
Reaction mixture
| Enzyme Solution: |
0.05 - 0.1 mg/ml for Zymolyase® - 20T |
1ml |
|
0.012 - 0.024 mg/ml for Zymolyase® - 100T |
1ml |
| Substrate: |
Brewer's yeast cell suspension (2 mg dry weight/ml) |
3 ml |
| Buffer: |
M/15 Phosphate buffer, pH 7.5 |
1 ml |
| Distilled Water: |
|
1 ml |
Procedure
After incubation for 2 hours at 25 deg. C with gentle shaking, A800 of the mixture is determined. As a reference, 1 ml of distilled water is used instead of enzyme solution.
Calculation
Percentage decrease in A800 = (A800 of reference - A800 of reaction mixture) x 100 / initial A800 of reference. 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® - 100T lyses 3 mg dry weight of brewer's yeast.
Precautions on Use
1) Avoid using nitrocellulose filters and use of material other than nitrocellulose, when sterilizing. Zymolyase® may be adsorbed on nitrocellulose membranes.
2) Zymolyase®, especially Zymolyase® -100T, may not be completely dissolved in buffers. Use Zymolyase® as suspension.
3) When sterilized, Zymolyase® is used in a concentration higher than 0.05%, prepare 2% Zymolyase® solution in buffers containing 5% glucose, filter the suspension and dilute the solution with the appropriate buffer.
References
1) Kaneko, T., Kitamura, K and Yamamoto,Y.: J. Gen. Appl.Microbiol. 15, 317 (1969)
2) Kitamura, K., Kaneko, T. and Yamamoto,Y.: Arch. Biochem. Biophys., 145, 402 (1971)
3) Kitamura, K., Kaneko, T. and Yamamoto,Y.: J. Hen. Appl.Microbiol., 18, 57 (1972)
4) Kitamura, K. and Yamamoto,Y.: Arch. Biochem. Biophys., 153, 403 (1972)
5) Kaneko, T., Kitamura, K. and Yamamoto,Y.: Agric. Biol. Chem., 37, 2295 (1973)
6) Kitamura, K., Kaneko, T. and Yamamoto,Y.: J. Gen Appl.Microbiol., 20, 323 (1974)
7) Kitamura, K. and Yamamoto,.: Agric. Biol. Chem., 45, 1761 (1981)
8) Katamura, K. and Tanabe, K.: Agric, Biol. Chem., 46, 553 (1982)
9) Katamura, K.: J. Ferment. Technol., 60, 257 (1982)
10) Kitamura, K.: Agric. Biol. Chem., 46, 963 (1982)
11) Kitamura, K.: Agric. Biol. Chem., 46, 2093 (1982)
12) Calza R. E., Schroeder A. L.: J. Ben.Microbiol., 129, 413 (1983)
13) Iizuka Masaru, Torii Yasuhiko,Yamamoto Takehiko: Agric. biol. Chem., 47 (12), 2267 (1983)
14) Shibata Nobuyuki, Kobayashi Hidemitsu, tojo Menehiro, Suzuki Shigeo: Arch. Biochem. Biophys., 251 (2), 697 (1986)
15) Iijima Y.,Yanagi S. O.: Agric. biol. CHem., 50 (7), 1855 (1986)
16) Herrero Enrique, Sanz Pascual. Sentandreu Rafael: J. Gen.Microbiol., 133 (10), 2895 (1987)
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