Enzymatic lysis

Lyse the cells in the presence of protease inhibitors either by enzymatic lysis (0.2 mg/ml lysozyme, 20 xg/ml DNAse, 1 mMmgCl2) or by mechanical lysis (Sonication, homogenization, repeated freeze/thaw) in 20mM sodium phosphate, 500 mM NaCI. Adjust the pH of the lysate to pH 7.4 using a dilute acid or base. 

Cell lysis Mechanical methods pressure shearing, ultrasonic disintegration, bead-mill homogenizers Nonmechanical methods enzymatic lysis, osmotic lysis, freezing and thawing, detergent-based lysis and electroporation... 

There are several methods of cell lysis (Table 8.2) [1,2], but there is none that works with cells of all biological origins. Each technique has its advantages and disadvantages, and the specific method of choice depends on the cell characteristics, the cell type, and the final application. A combination of more than one method may also be used. For example, enzymatic lysis uses specific enzymes to target the cell wall. However, to disrupt the cytoplasmic... 

Nonmechanical Methods of Cell Lysis Enzymatic Lysis... 

Enzymatic lysis Lysozyme/Triton X-100 < 100 ml Gram-positive bacteria... 

Structured and Simple Models of Enzymatic Lysis and Disruption of Yeast Cells... 

This paper presents two models of enzymatic lysis of yeast cells a simplified two-step model, accounting for protein release at cell lysis followed by proteolysis, and a more complex mechanistic model which describes the removal of the two layers of the yeast wall and the extrusion and rupture of the protoplast and organelles. 

On the right-hand side of equation 1, the initial term represents autolysis and the second term, enzymatic lysis. Equation 2 describes protein breakdown by product-degrading proteases. The first term on the right side stands for the protein released from lysing cells, and the second term, breakdown of the protein already in solution. Equation 3 shows that peptides are released from lysing yeast, but also arise from breakdown of longer proteins, P. 

Work continues in two areas purification of the lytic system to allow protease and glucanase levels to be controlled independently and investigation of the release of site-specific yeast enzymes and subcellular fractions by enzymatic lysis. 

Hunter, J.B. and Asenjo, J.A., "Kinetics of enzymatic lysis and disruption of yeast cells 1. Evaluation of two lytic systems with different properties." Biotechnol. Bioeng., 1986 (submitted for publication). 

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