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Chapter 20 : Practical Applications for Periplasmic Protein Accumulation
Category: Bacterial Pathogenesis; Microbial Genetics and Molecular Biology
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The biotechnology industry has exploited the unique properties of the periplasm to produce industrially relevant products. For efficient periplasmic protein accumulation to occur, events in the cytoplasm related to translation initiation and protein folding of the mature domain need to be coordinated. In this chapter the authors demonstrate that nucleotide changes in the translation initiation region of the gene profoundly influenced periplasmic protein accumulation. There are many documented examples of cytoplasmic proteins having positive effects on periplasmic protein accumulation of endogenous or recombinant proteins. A possible molecular explanation is that the cytoplasmic resident protein is preventing the preprotein from negative pathways, such as premature folding and inclusion body formation, and keeping the protein in an unfolded state and allowing efficient export. There are three possible fates for newly secreted proteins upon release into the periplasmic space. First, proteins can misfold and aggregate forming insoluble inclusion bodies. Second, proteins can be proteolyzed by endogenous periplasmic and membrane proteases where the protease domain of membrane proteases is located in the periplasm. Lastly, proteins can successfully fold into the correct conformation and exist as soluble, biologically active proteins. The lack of an identified chaperone system in the periplasm has been hindering the advancement of recombinant periplasmic protein accumulation. The combination of over-expression of periplasmic components participating in protein-folding pathways and the inclusion of chemical additives has been reported and shown to have positive benefits.
Key Concept Ranking
- Outer Membrane Proteins
Conceptual model of protein secretion in E. coli. The spigot represents the transcriptional induction of the recombinant gene and the funnels represent the major processes that are involved in periplas-mic protein accumulation. Each major process has several subprocesses that can be limiting, resulting in decreased yield of the intended protein. In this example there is a limitation in the cytoplasmic membrane secretion step, which could manifest as an increase in precursor protein accumulation.
Possible fates of proteins after secretion into the periplasm. Proteins have three potential fates in the periplasm: (i) aggregation and inclusion body formation, (ii) proteolysis, and (iii) folding to their proper conformation.