BIOTECHNOLOGY 61
Fig. 1. Electron microscopic image of B. megaterium (large cells) and E. coli (small cells). Photo courtesy of Prof. Dr. M. Rohde, TU Braunschweig, Germany.
Non-E. coli expression hosts represent a heyday for Gram- positive bacteria, says Arne Schulz.
Advances in bacterial hosts P
roteins are a group of organic molecules indispensable for life.
Tey are essential for the structure of all living organisms, and proteins also largely regulate the function of cells. Proteins include all enzymes and most hormones. Te main source of proteins is our daily food, deriving from plants and animals. Te human body breaks the proteins down to amino acids that then are used to form new proteins – in a process called protein synthesis. Dysregulation of this process is responsible for many diseases, for instance cancers and neurological disorders. Using bacteria as cellular factories and thus alternate sources of essential proteins is one of the major aims of gene expression.
A different aspect of artificial protein synthesis by means of bacteria is the production and supply of industrially relevant proteins. Bacillus subtilis is a major source of enzymes capable to degrade a variety of substrates. Popular examples are proteins
naturally secreted by Bacillus species into the growth medium, such as alkaline proteases as washing agent or amylases for the starch industry.
A major benefit of using bacteria as cellular factories for the production of proteins is that you can have them producing exactly the protein you want, and only a few steps are required from the genetically modified bacterium to the protein of interest in pure form.
Recent advances in preferred hosts Over the past decades, the intestinal Gram-negative bacterium Escherichia coli (commonly abbreviated to E. coli) has been the preferred host for artificial gene expression, mainly because it can be grown cost-effectively on widely available cheap substrates with high protein yields. However, its Gram-negative status, giving cause to problems related to the presence of endotoxins, renders it unable to efficiently secrete heterologous
proteins into the medium. Terefore, the availability of alternate hosts has become indispensable.
Tis gap could be filled by Gram- positive bacteria such as Bacillus megaterium, Bacillus subtilis and Lactococcus lactis. Tese bacterial hosts for gene expression and protein production are free of endotoxins, and efficiently working secretion pathways are present. In addition, Bacillus subtilis is classified by the FDA as GRAS (generally recognised as safe). A great variety of expression vectors for either bacterial host allows for the expression of a protein of interest.
Current state of play At present, about 60% of the commercially available enzymes are produced by Bacillus species, most notably Bacillus subtilis(1)
.
Vectors carrying signal peptides enable direct secretion of the protein into the growth medium, and different purification tags, for instance, His tag or Myc tag, facilitate easy purification(2) Work focusing on promoter
.
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