CORDIS - EU research results

Code-engineered new-to-nature microbial cell factories for novel and safety enhanced bio-production

Final Report Summary - METACODE (Code-engineered new-to-nature microbial cell factories for novel and safety enhanced bio-production)

Executive Summary:
The context of METACODE and its objectives
The questions and ideas about "what life really is" do not belong anymore to the exclusive domain of abstract academic, metaphysical and spiritual concepts. We are now interesting in this question also from the technological point of view. Latest by the discovery of the mechanisms of DNA biogenesis, scientists themselves finally changed their role from describing to analysing to finally start to engineering living systems.
Modern genetic engineers aim at building living systems, like true engineers build machines in order to establish technologies to create artificial biodiversity to produce virtually every imaginable medically or industrially interesting substance. In the coming post-industrial age we will face a strong growth of biology-based design with huge computer databases dealing with immense amounts of DNA sequences. Digitized information should enable us to recreate materials, living cells and organisms. Indeed, synthetic biology already consider cells (especially microbes) as robots and chemical machines equipped with DNA as software that defines the manufacture of proteins and other macromolecules that can be viewed as hardware. Changes in the genetic programme can, therefore, cause great variances in the produced substances. Driven with these ideas and visions, the METACODE consortium unified forces of eight partners from five European countries over last four years in order to generate artificial biodiversity.
The rationale behind the METACODE research philosophy is based in an assumption that artificial biodiversity will represent an important technology for the future, with living cells (i.e. mainly microbes) that function as small programmable production units. In the frame of the METACODE project, we made first steps towards the recruitment of novel bio-orthogonal chemistry (bio-compatible metathesis) with parallel genetic code engineering in microbial strains for mass production of tailored-to-fit protein/peptide-based antimicrobial products. This defines METACODE - metathesis and genetic code.
Beside the fact that we managed to make a significant scientific progress (see below), the METACODE consortium also succeeded to establish the new scientific discipline of Xenobiology (XB) , a marriage of chemical synthesis with synthetic biology, with potentials to build artificial biological systems and to address urgent technological problems. The foundational conference (XB1) was organized in Genoa and the second one in Dresden in the frame of METACODE efforts to raise awareness in the European general public about scientific, technological and societal impacts as well as the potentials of this new scientific discipline. We succeeded to gather scientists, engineers, designers, policy makers and other stakeholders to chart the paths toward an entirely novel biodiversity.

Project Context and Objectives:
Bacterial cells generated in METACODE consortia are equipped by orthogonal chemistries from the chemical synthetic laboratory which now become an integral part of the biochemistry of living cells. For example, we have demonstrated the in vivo applicability of the biotin-streptavidin technology to create an artificial metalloenzyme catalyzing olefin metathesis, a reaction absent from the natural enzyme’s repertoire. Doubtless, this will contribute to the future development and implementation of a set of bio-orthogonal tools to be utilized in new-to-nature metabolic pathways and beyond. Specifically, substantial progress made in the frame of METACODE consortia in the last 4 years include (a) evolution of the unique generalist acyl-tRNA ligases (b) metabolic syntheses of their substrates (c) long term evolution experiments that should enable bacterial adaptation (along with codon emancipation) with these substances, (d) important steps to establish and evolve an artificial metathesase in the periplasm of E. coli, and (e) the production of metabiotics, ribosomally synthetized peptides capable for modifications via metathesis reaction.
The success in the of engineered pyrrolysyl-tRNA synthetase that is capable of activating a metathesis-competent amino acid S-allylcysteine (Sac) for incorporation into recombinant proteins is further expanded by devising a synthetic pathway that enables in situ biosynthesis of this building block by re-designing the existing metabolic capabilities of Escherichia coli. In this way we succeeded in “teaching” E. coli cells to synthesize these building blocks on their own by using inexpensive starting substances. Our system engineering and long term evolution experiments reassign degenerate and rare (i.e. sense) codons of the genetic code. This breakthrough included the 'emancipation' of all 5797 AUA codons with and full reassignment of 20,899 UGG codons in the proteome of the bacterium Escherichia coli. These substantial breakthroughs represent a solid basis for the design of bacterial strains with altered genetic code that could lead to the creation of synthetic organisms (Fig.1). These synthetic cell factories already serve as useful platforms to evolve the catalytic performance of an artificial metalloenzymes and to enable the production of a novel generation of synthetic protein/peptide-based antimicrobial products.

Project Results:
see attached

Potential Impact:
see attached

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