Zymolyase® 20T, 1 g

Zymolyase^®, produced by a submerged culture of Arthrobacter luteus, has strong lytic activity against living cell walls of various strains of yeast cells. Hence, it is frequently used in order to produce protoplasts or spheroplasts.

The 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.

Other enzymatic activities contained: β-1,3-glucanase, protease, mannanase. Trace amounts of amylase, xylanase, phosphatase.

Stability: At 30 °C 70 % of the lytic activity is lost after 3 months.

Optimum temperature and pH: at pH 7,5 - 35 °C for lysis of viable yeast cells, at pH 6,5 - 45 °C for hydrolysis of yeast glucan.

Specificity (lytic spectrum): Ashbya, Candida, Debaryomyces, Eremothecium, Endomyces, Hansenula, Hanseniaspora, Kloeckera, Kluyveromyces, Lipomyces, Metschnikowia, Pichia, Pullularia, Torulopsis, Saccharomyces, Saccharomycopsis, Saccharomycodes, Schwanniomyces, etc.
Stock solution: As stock solution, a 2 to 10 % (20 or 100 mg/ml) Zymolyase^® solution may be prepared, solubilizing the enzyme in water, 10 mM sodiumphosphate buffer (pH 7,4) or 50 mM Tris-Cl (pH 7,5), respectively, according to the buffer system to be used downstream and containing 5 % glucose and 50 % glycerol each. The Zymolyase^® tends to not dissolve completely. Do not heat for solubilization, but rather use the Zymolyase^® as suspension. The suspension may be stored in aliquots at -20 °C.
Working solution: For use in the respective buffer system, dilute the stock solution to the working concentration of 2-5 mg/ml and optionally sterilize by filtration (0,2 µm). In most cases, however, Zymolyase^® is solubilized in freshly prepared working buffer (for instance rescue buffer 50 mM Tris-Cl, pH 7,5, 10 mM EDTA, 0,3 % β-Mercaptoethanol) in the required working concentration directly prior to use.
Sterile filtration: Avoid using nitrocellulose filters when sterilizing - Zymolyase^® may be adsorbed to nitrocellulose membranes.

One unit of lytic activity is defined as the enzyme amount causing a decrease of 30 % in absorbance at 800 nm using 6 mg Brewer’s yeast as substrate in Phosphate buffer (pH 7,5) at 25 °C. Lytic activity varies depending on the particular yeast strain, the growth stage of the yeast, and cultural conditions.

Zymolyase^® 20T is prepared by ammonium sulfate precipitatation.