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enGenes provides a high-quality protein expression platform for cost-effective biomolecule manufacture
enGenes Biotech provides high-quality protein expression and process development services to enable cost-effective manufacturing of biomolecules, including recombinant protein production, secretory production of proteins and plasmids DNA production. enGenes can provide milligrams and even grams quantities of proteins and various plasmid DNA for research purposes and also specializes in upstream and downstream processes development for the large-scale production of research-grade plasmids DNA.
enGenes is an expert in the use of the T7 expression system to clone recombinant DNA using strains of Escherichia coli (E. coli) as an optimal system for expressing recombinant proteins. Expression of recombinant proteins under the control of the T7 promoter is up to eight times faster than basic E. coli expression under the control of RNA polymerase.
In the T7 expression system, an expression vector such as pET is adapted to bring together a T7 promoter with a gene of interest that the promoter will control during downstream processing (DSP). The combined expression vector is transformed into an E. coli strain. The E. coli cell’s chromosomal gene can be expressed to produce T7 RNA polymerase.
Transcription is initiated when an inducer binds to the lac repressor to prevent it from inhibiting the E. coli expression of the T7 gene. T7 RNA polymerase binds to the T7 promoter on the expression vector to transcribe to its downstream gene of interest. To stimulate production and fermentation processes, the inducer reagent IPTG can be added to prevent the lac repressor from inhibiting gene expression. enGenes Biotech has extensive know-how and proprietary technologies to avoid ‘leaky expression’ and to prevent plasmid instability.
What is an expression vector?
Expression vectors are the basic tools in biotechnology for the production of proteins. An expression vector, otherwise known as an expression construct, is usually a plasmid or virus designed for gene expression in cells. The vector is used to introduce a specific gene into a target cell, using the cell’s mechanism for protein synthesis to produce the protein encoded by the gene. A particularly well-known example of an expression vector is the production of insulin, which is used for medical treatments of diabetes.
enGenes combines in-depth experience in cutting-edge genetic engineering with broad expertise in bioprocess engineering to deliver best-in-class protein expression systems. Based on a strong understanding of the requirements of the biotech fermentation industry and extensive knowledge and method portfolio, enGenes delivers improved host cell lines, plasmid expression vectors and optimized production processes to maximize bio-process efficacy.
Leading-edge expression technology
enGenes Biotech has developed leading-edge platforms for the customized production of recombinant proteins and plasmids DNA for AAV production that can frequently prove more cost-effective than a competing pharmaceutical contract development and manufacturing organization (CDMO) can offer.
These enGenes microbial E. coli expression solutions are based on its unique and proprietary enGenes -eX-press bacterial cell line technology platform, enhanced by patented manufacturing processes to leverage the basic -eX-Press growth decoupled advantage. The net results are substantially higher soluble product yields and easier downstream processing (DSP).
enGenes-eX-press forms a next-generation E. coli expression platform that allows for scalable production of difficult-to-express or even “toxic” recombinant proteins and high-quality supercoiled plasmids DNA at a fraction of previous cost and within short time frames to make unique biotech products commercially viable. Patents are pending on -eX-press technology as a growth-decoupled microbial system to produce proteins and plasmid DNA.
Harnessing E. coli molecular capabilities
Bioprocesses harnessing the properties of cells and enzymes through tools such as recombinant DNA and tissue culture presents multiple challenges, particularly in downstream processing and scale-up to industrial viability.
Manufacture of typical recombinant DNA-derived products will involve up to 20 steps, with the first half of the process devoted to genetic manipulation of the host organism to establish levels of expression available and the stability of the constructed strain of E. coli, defined as characteristics of the recombinant cells as a function of the E. coli culture environment.
Once viable E. coli culture conditions for production are established, process development will involve using a benchtop bioreactor to establish ideal process settings, such as acidity, oxygen levels, temperature, etc.
The unique enGenes-eX-press™ proprietary technology platform for outstanding recombinant protein production in E. coli uses an advanced growth decoupled mechanism platform that allows customers to achieve customized solutions for the cost-effective production of recombinant proteins in microbial expression systems.
The enGenes -eX-press platform optimizes the use of E. coli as the host for proteins production based on a genetically modified host cell based on BL21 (DE3) that allows for the introduction of standard expression vectors (with T7 promoter) with no need for cumbersome adaptations.
It uses an advanced growth decoupled mechanism platform that allows customers to achieve customized solutions for the cost-effective production of recombinant proteins and plasmid DNA production in microbial expression systems.
The -eX-press fermentation process can be implemented with standard fermentation chemistry equipment.
Reprogramming of the host cell is performed by co-expression of a bacteriophage-derived peptide that stops cell division and host mRNA production and at the same time modulates the host cell metabolism for improved soluble, high-level protein production.
Adaptable fermentation method
-eX-press is a versatile technology that can be applied in many ways in conjunction with other enGenes products to produce proteins, enzymes, bioconjugates, metabolites, small molecules, and plasmid DNA for gene therapy or AAV production.
These related technologies include:
-eX-tra for extracellular secretion of recombinant proteins
-eX-pand for expansion of genetic code and incorporation of non-canonical amino acids
-eX-cite for continuous production of biomolecules
-eX-cess for pDNA upstream and downstream processing and growth-decoupled pDNA production
-eX-change for antibiotic resistance market free protein of biomolecules.
Used in combination, this suite of technologies has the potential to achieve more than five-fold increases in yields.
enGenes -eX-press optimized protein expression system
enGenes Biotech has developed Intellectual Property (IP) and has patents pending on its enGenes-eX-press™ technology which forms a growth-decoupled microbial protein production system for optimized protein expression in E. coli.
This prokaryotic protein expression system allows enGenes Biotech to offer a range of highly relevant products and services to companies seeking to develop cost-effective manufacturing processes for recombinant proteins.
enGenes is also developing IP in related research fields, such as microbial screening of high-producer clones, improved plasmid vector expression systems and antibiotic resistance-free plasmid selection for recombinant protein productions.
Optimized protein expression in insect cells
enGenes also offers optimized protein expression using the Baculovirus Expression Vector System (BEVS) as a complementary service for the production of functional soluble proteins that are not easily expressed in E. coli. BEVS is universally recognized as a powerful and versatile system for the production of high-quality proteins.
Protein expression using BEVS is offered for leading insect cell lines such as Sf9, High Five and Tnao38.
BEVS service options include recombinant baculovirus range transfection and selection, amplification of high-titer virus stock, small-scale time course expression test, large-scale expression and purification, through to the establishment of the purification protocol and final purification.
enGenes protein expression service
Based on this extensive knowledge and fermentation method portfolio, enGenes can deliver improved host cell lines, plasmid expression vectors and optimized production processes to maximize bioprocess efficacy.
enGenes -eX-press forms the key building block in a modular system of customer services for gene expression and optimized protein production in E. coli and other prokaryotes, such as Bacillus subtilis.
The enGenes optimised protein expression service is divided into a series of steps covering all specific demands along the line from gene to protein.
These steps can be performed either individually or in combination, according to customer preferences and include:
enGenes service options include:
- Cloning / Vector optimization
- Generation of expression coli expression strain / Host cell engineering
- High-Throughput Screening of Expression Clones
- Feasibility study at lab scale in fed-batch mode
- Development of Downstream Processing routes
- Scale-Up and Tech-Transfer to CMOs
Along with the regular project updates, clients also receive step-by-step progress reports with detailed documentation to comply with their internal quality systems.
Contract manufacturing advantages
enGenes operates in the role of a contract research organisation (CRO) that can partner with chosen contract manufacturing organisation (CMO) or contract development and manufacturing organization (CDMO).
Even in advance of the project research phase, enGenes can liaise with the customer’s chosen CDMO or CMO manufacturing organisation to ensure smooth tech transfer with upstream and downstream pre-developed and precise yield estimates for the fed-batch process of fermentation to deliver the best value for the client.