14IBCAT1FEASIBILITY - Engineering a Nano-factory for Peptide Synthesis

Lead Research Organisation: University of Bristol
Department Name: Biochemistry

Abstract

Peptides are short chains of simple chemical building blocks called amino acids. They are involved in numerous key biological processes including acting as toxins, pigments, drugs and hormones. They also control many of the most important cellular functions in animals, plants and man, and are used as tools in scientific research. There is considerable worldwide interest in developing new methods of producing peptides in sufficient quantity and of sufficient quality for use as pharmaceuticals, agrochemicals or research tools. The current favoured method for manufacturing peptides involves using chemical agents to fuse together the amino acid building blocks that form them. This approach is time-consuming, expensive, generates toxic waste products, and cannot be applied to many unusual but valuable peptides. In this project we will develop a new method of producing peptides which is not limited by these major problems. We will use an engineered peptide 'nano-factory', which when introduced into bacteria allows them to produce significant quantities of high value 'difficult' peptides without any of the problems or complexities associated with chemical synthesis.

Technical Summary

The global custom research peptide manufacturing market is currently valued at approximately £800M per annum. The future challenges facing this industry revolve around developing tractable routes to producing increasingly complex peptides, increasing peptide synthesis capability, and a requirement for new tools that facilitate access to historically intractable high value targets, e.g. amyloids, anti-microbial peptides, hydrophobic peptides and isotopically-labelled peptides. We have developed a bio-inspired nano-scale device, based on a ubiquitous bacterial chaperone, that sequesters and stabilises peptides during their biosynthesis, protecting them from proteolytic action and maintaining them in a fully folded and, where appropriate, a functional state. The objectives of this proposal are (i) to rationally re-engineer our prototype device to produce functionally-enhanced variants with broader utility, (ii) to integrate this technology into optimised microbial and cell-free expression platforms, and (iii) develop methods for efficient extraction and purification of the peptides following recombinant production, with a view to optimising peptide purity. Further preliminary investigations will take place to scale-up production with a view to the development of rapid methodologies for commercial production of pure peptides. The development of a ubiquitous method for recombinant protein production, even of peptides that are difficult to synthesise by traditional solid-state chemical methods, will provide great benefits.

Planned Impact

As described in proposal submitted to IUK.

Publications

10 25 50
 
Description We have developed a platform technology for the recombinant production of peptides that have yields as good as those produced by the more common solid-phase peptide synthesis and have a greater degree of purity, especially for longer peptides (> 30 amino acids in length). In addition we are able to produce peptides that are difficult to synthesise by conventional solid-phase chemistry (e.g. beta-amyloid 1-42). The process also has similar economic costs to solid-phase synthesis but, in addition to purity, the process also produces approximately 98% less environmentally damaging waste products (e.g. organic solvents). We are currently using a BBSRC Impact Acceleration Account award to employ a business consultant to assist us with the development of a route for commercialisation of our production technology.
Exploitation Route We are already taking steps to commercialise our technology with the intention of potentially licencing the basic technology while also developing it further for the production of much larger quantities of peptide and also randomised peptide libraries for screening. Since many new therapeutic compounds are peptides, as well some cosmetic products and synthetic biology materials, we envisage a large market that requires the production of pure peptides that are difficult to synthesise using solid-phase chemistry and also in much higher quantities than are currently available.
Sectors Agriculture, Food and Drink,Chemicals,Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology

 
Description The platform technology developed in this grant formed the basis of the spin-out company Zentraxa Ltd. Zentraxa is using this platform to produce a prototype peptide-based bioadhesive molecules as the basis of a novel, transparent, waterproof adhesive for use by the Ministry of Defence. This will be developed further to scale-up production and produce the correct formulation for use in applications. Zentraxa Ltd are also developing other bioadhesives and biomaterials to be produced using this platform, including a medical adhesive for use in surgical procedures. Zentraxa Ltd went through a first round of investment in May 2019 and were also awarded a fresh contract from DSTL as well as commercial engagement with Qinetiq.
First Year Of Impact 2017
Sector Aerospace, Defence and Marine,Government, Democracy and Justice,Manufacturing, including Industrial Biotechology
Impact Types Economic

 
Description Impact Acceleration Account
Amount £18,820 (GBP)
Organisation Engineering and Physical Sciences Research Council (EPSRC) 
Sector Public
Country United Kingdom
Start 08/2017 
End 07/2018
 
Description Impact Acceleration Account
Amount £10,000 (GBP)
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 01/2016 
End 06/2016
 
Description Materials and Manufacturing Round 3
Amount £100,000 (GBP)
Organisation Innovate UK 
Sector Public
Country United Kingdom
Start 02/2018 
End 01/2019
 
Description Pathfinder
Amount £11,972 (GBP)
Funding ID BB/P022839/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 12/2016 
End 03/2017
 
Title Recombinant method for the synthesis of peptides 
Description While solid-phase peptide synthesis (SPSS) is generally the method of choice for peptide synthesis it suffers from some inherent drawbacks. Peptides that are very hydrophobic, and hence sparsely soluble in aqueous conditions, are often difficult to manufacture, as are peptides that have highly repeating amino acid sequences. We have engineered a recombinant system to produce such peptides. Protein synthesis from the bacterial ribosome is more efficient and accurate than SPSS and is not limited by the sequence of amino acids. Our recombinant system produces peptides in a tagged and insoluble form that is targeted to intracellular inclusion bodies. These can be rapidly solubilised and purified, and the tag proteolytically-cleaved to produce high yields of peptide. These can then be further purified, if necessary, using conventional reverse-phase HPLC. 
Type Of Material Technology assay or reagent 
Year Produced 2017 
Provided To Others? No  
Impact The technology can be easily scaled up for industrial scale production using large volume bacterial fermentation facilities. One other advantage of this methodology is that it reduces the amount of organic solvent waste by greater than 90%, producing only biodegradable waste. This has the potential to reduce the environmental impact of industrial processes requiring large-scale peptide synthesis. It is currently being used by a University of Bristol spin-out company, Zentraxa Ltd, for the synthesis of peptide-based bioadhesive molecules. 
 
Description DSTL 
Organisation Defence Science & Technology Laboratory (DSTL)
Country United Kingdom 
Sector Public 
PI Contribution The work partially funded by this grant was used to develop a platform technology for producing peptide molecules that are difficult to synthesise by conventional solid-phase synthetic chemistry techniques. Our spin-out company, Zentraxa Ltd, is using this technology in the development of water-resistant bioadhesives, based on the adhesive chemistry of the mussel foot protein. These peptides or protein fragments are impossible to synthesise by conventional means. This is being done as a collaborative partnership with DSTL.
Collaborator Contribution DSTL initially provided funding of £100K to develop the application of our peptide production platform technology to the problem of water-resistant bioadhesives. This funding allowed us to provide a proof-of-principle adhesive molecule and begin the process of optimisation.
Impact Zentraxa has produced water-resistant bioadhesive molecules using their proprietary technology, Zentide, to the satisfaction of DSTL. DSTL have recently agreed to fund further R&D, also in association with Qinetiq, to further advance this technology with the aim of establishing a solution for a specific military problem, but one that is also readily transferable to other non-military applications. The bioadhesive molecules are currently the subject of a patent application that is being prepared.
Start Year 2017
 
Company Name Zentraxa Limited 
Description Zentraxa's proprietary technological platform, Zentide, combined with extensive technical know-how pushes the boundaries of what can be achieved with synthetic biopolymers. The company specialises in the design, production and testing of complex novel peptides and, as a result of our proprietary peptide biosynthesis platform, has the ability to circumvent previously existing bio-design limits imposed by conventional peptide synthesis, with one universal process. 
Year Established 2017 
Impact The company has designed and produced a prototype transparent, water-proof bioadhesive for DSTL, our launch customer. This has currently completed testing and will enter a second phase of R&D to scale-up production and develop the correct formulation for use. Zentraxa is also extending these designs for use in a clinical setting, to replace cyanoacrylate-based adhesives that are currently used in surgical procedures. The company has recently been through its first investment round raising £0.5M of investment from angel investors. The company has also moved out of the University of Bristol and has established its own base of operations at the UnitDX incubator in the centre of Bristol.
Website http://www.zentraxa.com