T7 lysome come from pLysS cause lysis?

davesmith at bioch.tamu.edu davesmith at bioch.tamu.edu
Tue Jul 1 00:41:15 EST 1997


e_yckim at cais.kaist.ac.kr (Yeon Chul Kim) wrote:

>Hi!!

>We performed fed-batch fermentation using pLysS containg E. coli.
>After IPTG induction, we acheived ca 90 g/L of DCW.  But a lots of
>cells are lysed.   I think cell lysis is common problem in high cell
>density.  But, I also wonder the T7 lysome produced by pLysS cause
>lysis.  In some reference, relatively high levels of T7 lysome can be
>tolerated by E. coli, because the protein is unable to pass through
>the inner membrane to reach the peptidoglycan layer.  But,  I think,
>in high cell density, cells are not in normal conditions.  So, the T7
>lysome enhance the cell lysis.
>Do you have any comment?
>Or Do you have some reference about pLysS plasmid (especially, amount
>of T7 lysome produced, effect on cell lysis.)


You wouldn't by chance be over-expressing an integral membrane protein
or secreted protein, would you?

In order to get lysozyme  past the inner membrane, the membrane must
be abnormally perturbed.  I don't think induction of T7RNAP alone will
result in this sort of  membrane perturbation.  We work with the phage
protein responsible for "letting lysozyme out" at the time of lysis
and use IPTG to induce in our over-expression system.  In the vector
controls we do not see lysis.

Dave.




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