Hi,
If I express membrane protein of Mycobacterium in E.Coli cells, is it necessary to have signal peptide ? How does E. coli machinery sends it to membrane? Is there is any specific pathway like signal recognition particle (SRP)?
Any other thing to be careful when I want to express it in membrane?
Any links of paper in this regard will be highly helpful.
If it is a membrane protein, then a signal peptide will not only locate the protein to the E.coli membrane (especially if you engineer it to the signal peptide sequence typical for E. coli) but will also prevent the membrane protein accumulating in the cytoplasm which may prove lethal for the cell. I suggest you aim to ensure it is located to the E. coli membrane.
The following articles on signal peptides may be useful:
1. Nielsen, H., Engelbrecht, J., Brunak, S & von Heijne, G. (1997)
Identification of prokaryotic and eukaryotic signal peptides and
prediction of their cleavage sites. Protein Engineering
vol.10 no.1 pp.1–6, 1997
2. By same authors: The signal peptide. J. Membrane Biology 115: 195-201.
Our lab has also published work on the expression of membrane proteins in E.coli:
Azam, A.A., Kinder, J.M., Khan, G.N., Alase, A., Ma, P., Liu, Y., Ault, J.R., Henderson, P.J.F., Chowdhry, B., Alexander, B.D., Harding, S.E. & Phillips-Jones, M.K. (2018) Production of membrane proteins for characterisation of their pheromone-sensing and antimicrobial resistance functions Eur. Biophys. J in press (attached)
and
Hussain, R., Harding, S.E., Hughes, C.S., Ma, P., Patching, S.G., Edara, S., Siligardi, G., Henderson, P.J.F. & Phillips-Jones, M.K. (2016) Purification of bacterial membrane sensor kinases and biophysical methods for determination of their ligand and inhibitor interactions. Biochem. Soc. Trans. 44: 810-823.
Good luck
