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Lysozyme attacks peptidoglycan, which are found in Gram-positive bacteria. I know someone who uses lysozyme to lyse Escherichia coli, which is Gram-negative. How is that possible? Can lysozyme lyse all bacteria, independently of their Gram status? Does anybody have information on how long I should leave lysozyme in a bacterial solution for it to be effective?

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Gram-negative bacteria also have peptidoglycan which can be degraded by lysozyme. However they have an outer membrane lying outside the peptidoglycan layer and this will block lysozyme action.

Because of this if you intend to treat Gram-negative bacteria with lysozyme it is necessary to add something that will disrupt the outer membrane—the usual choice is EDTA since removal of magnesium ions by chelation will destabilise the outer membrane.

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  • $\begingroup$ Do you know what concentration of EDTA and lisozyme would be ideal for killing the bacteria? $\endgroup$ – charlesdarwin Feb 4 '18 at 23:45
  • $\begingroup$ I've heard that lysozyme can, even in vivo, attack gram-negative bacteria through some alternative mechanism possibly unrelated to attacking peptidoglycan. $\endgroup$ – forest Feb 5 '18 at 5:04
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    $\begingroup$ @charlesdarwin Published protocols tend to focus on using EDTA/lysozyme treatment to prepare spheroplasts rather than killing. See ncbi.nlm.nih.gov/pmc/articles/PMC4223228 for an example. Note that the authors add sucrose to prevent the cells from lysing i.e. as an osmoprotectant. Their report indicates that rather short treatments (5 min) are enough to form spheroplasts: in the absence of sucrose these cells would have lysed. $\endgroup$ – Alan Boyd Feb 7 '18 at 11:27
  • $\begingroup$ Do you know what they mean by '125 mM EDTA-NaOH'? Is it 125 mM EDTA and 125 mM NaOH? $\endgroup$ – charlesdarwin Feb 12 '18 at 0:18
  • $\begingroup$ Also, do you think that the bacterial proteins will be denatured after the lysis of the bacterial cell membrane? $\endgroup$ – charlesdarwin Feb 12 '18 at 0:32

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