Team:Warsaw/Project/cytoplasm

Secretion of proteins into the cytoplasm

Theoretical basis

Bacterial systems of protein secretion

Free living bacteria make use of a diverse spectrum of transport and secretion systems. In gram negative bacteria type I secretion systems are very common e.g. the well characterized hemolysin secretion system. Hemolysin is responsible for the pathogenicity of some E. coli strains. Obtaining a fusion of a protein of interest with the hemolysin C-terminus results in the secretion of the protein into the growth medium[6] The same method can be used to secret bacterial proteins into the cytoplasm. This has already found its application in the new generation of vaccines[5].

Details

The cytoplasmic operon.

Fig 4. Overview of endosomal detection operon. It is composed of YFP – Yellow Fluorescent Protein, tetR – repressor of a tetracycline resistance operon, cro box regulatory sequence from λ phage and genes specific for a desired function of bacterial strain. In this case p53 – an apoptosis inducer.

Cytoplasmic operon, like the invasion operon, is a bistable switch. The regulatory elements are: a synthetic promoter inhibited by TetR, promoter inhibited by araC, the araC protein (inactivated by L-arabinose) and the TetR protein (repressor of the tetracycline resistance gene inactivated by tetracycline). [18] This switch controls genes expressed in the cytoplasm. Cro box confines the gene expression to the cytoplasm only. The cro protein bound to it stops the transcription while the endosome detection operon is expressed. The yellow florescence protein is expressed to allow for the detection of the cytoplasmic operon activity. The set of genes in the cytoplasmic operon depends on the desired function of a specific bacterial strain. In the case of an antitumor therapy this could be the p53 protein with mitochondrial leader sequence or the Bax protein. In the strain capable of conjugation with mitochondria cytoplasmic operon will control genes responsible for the conjugation process.