Inclusion bodies (bacteria)

In cells under an electron microscope most often, inclusion bodies look like large dark clusters ^{[1] [2]} . Inclusions isolated from body cells are amorphous spherical or rod-shaped formations with a diameter of 0.2 μm to 1.2 μm ^{[3] [4] [5]} .

The basis of inclusion bodies is a protein overexpressed in bacteria. According to some studies, the proportion of β-structures is increased in inclusion bodies and, therefore, many inclusion bodies are amyloid ^[6] . Most often, in addition to the main protein, inclusion bodies contain chaperons DnaK and GroEL, as well as two specialized proteins IbpA (Inclusion body protein A) and IbpB (Inclusion body protein B), found mainly in inclusion bodies ^{[7] [8]} . Chaperones DnaK is located on the surface, where it, together with ClpB, is involved in the destruction of the protein aggregate and protein refolding, while GroEL is inside the inclusion bodies ^[9] . Inclusion bodies may also contain additional proteins, depending on which particular protein is expressed. Thus, the inclusion bodies of the human main fibroblast growth factor hFGF-2 additionally contained DnaK chaperone, as well as translation factor EF-Tu and metabolic enzymes dihydrolipoamide dehydrogenase LpdA, tryptophanase TnaA, tagalose-1,6-bisphosphate aldolase GatY ^[10] .

It has been shown that protein aggregation in inclusion bodies is a reversible process. If protein synthesis ceases, inclusion bodies gradually disappear and a completely folded protein appears in the cytoplasm ^[11] . This process takes place with the participation of DnaK and ClpB chaperones and the active use of ATP hydrolysis energy ^{[12] [13]} . In the process of disintegration of inclusion bodies, IbpA and IbpB proteins and Lon and ClpP proteases can also be involved ^[14] .

Inclusion bodies contain relatively pure expressed protein and are relatively easily secreted. The only problem is the subsequent protein folding (re-folding). Therefore, there are even special expression systems that intentionally direct the protein into inclusion bodies. In particular, the combination of the expressed protein with proteins such as TrpLE, PurF, PagP, ketosteroid isomerase, leads to the formation of inclusion bodies of the desired purity ^[15] . The following methods of refolding are distinguished: dilution of a protein solution, dialysis , chromatographic refolding, and the use of high hydrostatic pressure ^[16] .