Sophisticated Cloning, Fermentation, and Purification Technologies for an Enhanced Therapeutic Protein Production: A Review

Abstract

The protein productions strategies are crucial towards the development of application based research and elucidating the novel purification strategies for industrial production. Currently, there are few innovative avenues are studies for cloning, upstream, and purification through efficient bioprocess development. Such strategies are beneficial for industries as well as proven to be vital for effectual therapeutic protein development. Though, these techniques are well documented, but, there is scope of addition to current knowledge with novel and new approaches and it will pave new avenues in production of recombinant microbial and non-microbial proteins including secondary metabolites. In this review, we have focussed on the recent development in clone selection, various modern fermentation and purification technologies and future directions in these emerging areas. Moreover, we have also highlighted notable perspectives and challenges involved in the bioengineering of such proteins, including quality by design, gene editing and pioneering ideas. The biopharmaceutical industries continue to shift towards more flexible, automated platforms and economical product development, which in turn can help in developing the cost effective processes and affordable drug development for a large community.

Progress in biopharmaceutical development

“…Additional favorable microbial recombinant protein production systems are the eukaryotic microorganisms S. cerevisiae and P. pastoris . Both of these hosts are capable of producing recombinant proteins with proper folding and posttranslational modifications .…”

“…The S. cerevisiae yeast expression system is frequently used due to their rapid growth in protein‐free media and ability to secrete the product extracellularly. However, posttranslational modifications that occur within the cells often lead to the production of undesired hypermannosylation , which can result in altered protein binding activity, and potentially yield an altered immunogenic response in therapeutic applications. In P. pastoris , oligosaccharides have much shorter chain lengths and the strain has been reported to produce complex, terminally sialylated or “humanized” glycoproteins .…”

The “Three Cs” of Novel Antibiotic Discovery and Production through Synthetic Biology: Biosynthetic Gene Clusters, Heterologous Chassis, and Synthetic Microbial Consortia

“…Recent work has suggested that this is not necessarily always the case, with the identification and introduction of a key regulatory element (the nonclustered LuxR‐type quorum‐sensing receptor, PviR, from P. luteoviolacea 2ta16) into E. coli increasing pathway transcription and production of violacein—a proof of concept compound—by ≈60‐fold . This work makes a case for identification of other regulatory elements which may increase the feasibility of E. coli in use for heterologous expression of antibiotic biosynthetic gene clusters, although it does not account for the level of metabolic burden associated with large heterologous proteins expressed in E. coli , as well as the organism's poor capacity for secretion …”

Three-dimensional culture of chicken primordial germ cells in chemically defined media containing the functional polymer FP003

“…Escherichia coli has been used to produce recombinant proteins because it can be easily cultured and is amenable to genetic modification. However, the production of recombinant proteins using this system is hampered by a lack of post-translational modifications (PTMs) and the risk of endotoxin contamination [5]. Recombinant proteins are also frequently produced in yeasts, such as Saccharomyces cerevisiae and Pichia pastoris .…”

“…Recombinant proteins are also frequently produced in yeasts, such as Saccharomyces cerevisiae and Pichia pastoris . Although yeast cells can be easily and inexpensively cultured, this approach is restricted by the limited number of yeast vectors and promoters as well as the lack of PTMs observed in human cells [5, 6]. Production of recombinant proteins in animal cells is a promising alternative with various clinical applications.…”