Transposition of animal and plant biosynthetic pathways in yeast cells:
from successes to challenges

Denis Pompon

The feasibility of designing artificial metabolic pathways transposing in microbes higher eukaryotes secondary metabolisms targeting important industrial molecules was demonstrated by several recent success stories. However, such transpositions still raised numbers of challenging questions needing to be mastered to make industrial successes of white biotechnology approaches not exceptions but a rule. The problem of metabolic transposition significantly differs from biosynthetic retargeting or optimization in a homologous context. Difficulties particularly arise from the need to couple the artificial pathway with host metabolism; to rebuild the compartmentalisation and transport mechanisms and to control the interaction of the newly produced metabolites with endogenous activities. In the absence of natural mechanisms of regulation, keeping in line an artificial pathway toward the wanted product is another difficulty. Furthermore, dealing with the necessary balance between host viability, genetic stability of the artificial pathway and productivity is also critical. The identification of suitable gene collections to be assembled for building synthetic metabolisms was a significant problem in the pre-genomic era. The current availability of large collections of annotated sequences and the development of sophisticated protein engineering technologies involving the association of rational (structure based) design with combinatorial optimisation methods, like directed evolution, has now greatly simplified this task.

These features will be illustrated by the transfer, in baker yeast, of more or less complex human metabolic pathways like the metabolism of drugs, the reconstruction of cholesterol biosynthesis or the self-sufficient biosynthesis of commercially important steroid hormones. Such transfers open access to a large range of applications in the field of drug safety and discovery as well as in the biotechnological production of active principles, constituting competitive and durable alternatives to chemical processes.

However, significant challenges remain to be addressed to make the design of artificial pathways cost effective and rapid enough to meet industry requirement. Most of current works remain fairly empirical and time consuming due to a wide use of trial and error approaches and step by step (serial) reconstruction strategies. Global optimisations of such artificial pathways are thus rather inefficient as occurring very late in the reconstruction process. Large gain in time and efficiency would be in contrast expected if combinatorial instead of serial approaches could be designed. Such parallel approach would in fact mimic at a metabolic network level what is currently done when addressing protein engineering through directed evolution processes. However, to date, tools required to develop such combinatorial approaches of metabolic engineering are still very limited. Host like yeast, which naturally exhibits efficient recombination machineries, could be of particular interest to develop such a strategy. Combinatorial approaches in addition to greatly accelerate transposition of specific metabolisms would be also expected to bring innovative solutions to constitute collection of microorganisms synthesising collections of active and novel compounds of interest.

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