Lysozyme fusion protein-bacteria

From: R.Srinivasan
Date: 22 February 2012 19:25

Dear All,

       We are working with proteins with a lot of surface hydrophobicity and hence solubility is a big issue. We so far have tried expressing them as fusion proteins.The strategy has yielded soluble protein but most of the protein elutes in the void volume on a gel-filtration colum.

      This brings me to my question, "Are there vectors which have an inactive lysozyme fusion tag, which could be used for expression in E.coli?".

Many thanks to all in anticipation,

Vasan

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From: Pius Padayatti

Although this is not the answer to your question,
Bryan Matthews papers show
In T4 lysozyme A mutation at A73 to AAA is shown to make extensive
core-repacking in T4lysozyme that makes the
enzyme inactive.
Is the issue here is to use lysozyme for bacterial cell breakage while
your fusion
protein remain inactive? Bacterial cells can be otherwise disrupted easily

So like i mentioned Poteete and co workers (Rennell eta al., 1991,
systematic mutation
of bacteriophage t4 lysozyme JMB 222:67-87)  mutated 164 sites with 13
different amino acids
50 % of the sites all 13 susb. left the protein like wild type.
So the A73 mutation mentioned above can be incorporated into your constructs?


padayatti