Recent advances in synthetic biology for engineering isoprenoid production in yeast Claudia E Vickers 1,2 , Thomas C Williams 3 , Bingyin Peng 1 and Joel Cherry 4 Isoprenoids (terpenes/terpenoids) have many useful industrial applications, but are often not produced at industrially viable level in their natural sources. Synthetic biology approaches have been used extensively to reconstruct metabolic pathways in tractable microbial hosts such as yeast and re-engineer pathways and networks to increase yields. Here we review recent advances in this field, focusing on central carbon metabolism engineering to increase precursor supply, re- directing carbon flux for production of C10, C15, or C20 isoprenoids, and chemical decoration of high value diterpenoids (C20). We also overview other novel synthetic biology strategies that have potential utility in yeast isoprenoid pathway engineering. Finally, we address the question of what is required in the future to move the field forwards. Addresses 1 Australian Institute for Bioengineering and Nanotechnology, The University of Queensland, St Lucia, QLD, Australia 2 Commonwealth Research Organization (CSIRO), Land and Water, Ecosciences Precinct, 41 Boggo Road, Dutton Park 4102, Queensland, Australia 3 Department of Chemistry and Biomolecular Sciences, Macquarie University, NSW, Australia 4 Amyris, Emeryville, CA, USA Corresponding author: Vickers, Claudia E (c.vickers@uq.edu.au) Current Opinion in Chemical Biology 2017, 40:47–56 This review comes from a themed issue on Synthetic biology Edited by Tom Ellis and Michael Jewett http://dx.doi.org/10.1016/j.cbpa.2017.05.017 1367-5931/# 2017 Elsevier Ltd. All rights reserved. Introduction The isoprenoid (terpene/terpenoid) family of natural products is home to a large variety of industrially-useful compounds. These range from relatively low-volume, high-value products (e.g. pharmaceuticals) through to high-volume, low value products such as biofuels and bulk industrial chemicals (Figure 1). Regardless of the market size, significant engineering is generally required to achieve sufficient rates, titres, and yields for industrial applications. Isoprenoid biosynthetic pathways have therefore been the subject of extensive research, and have become model systems for development of novel synthetic biology tools aimed at controlling pathway flux for metabolic engineering applications. Of the various production organisms available, yeast has emerged as the most successful due to a wide range of advantages, including: ease of manipulation; depth of genetic and physiological characterisation; availability of engineering tools; high sugar catabolic rate and relatively fast growth rate; relatively high native isoprenoid pathway flux, good capacity to engineer improved isoprenoid pathway flux and to introduce complex functional modifications of products; GRAS status; and amenability to industrial bioprocess conditions. The rate/titre/yield challenge is obviously most signifi- cant for commodity products [1]. Global energy security concerns coupled with environmental concerns around traditional petrochemical products have focussed atten- tion on biofuels in the commodity market. The availabil- ity of potential isoprenoid-based drop-in components for a range of fuels from diesel to jet has necessitated sub- stantial innovation in pathway engineering. While we are yet to meet the production efficiencies required for cost- effective production of isoprenoid biofuels, these innova- tions have provided platform technologies that are now applied to production systems for lower-volume, higher- value products — including mid-range industrial chemi- cals such as the cosmetic ingredient squalene, liquid farnesene rubber, vitamin precursors, fragrances, and (relatively) high-value pharmaceuticals such as the an- ti-malarial artemisinin [2,3]. Yeast operates a native mevalonate (MVA) pathway for isoprenoid production (Figure 2). The basic engineering steps for improved pathway flux have been well defined (reviewed in [4–7]). The initial step is usually introduc- tion of a specific product pathway; this serves the dual purpose of providing a metabolic route for production and providing a carbon sink to prevent build-up of prenyl phosphate intermediates, which mediate feedback regu- lation and may cause toxicity at high intracellular levels [8,9]. MVA pathway flux is then bolstered by expressing native or heterologous MVA pathway genes and further increased by engineering central carbon metabolism. Competing pathways are down-regulated, accumulation of pathway intermediates (which may cause toxicity or simply represent inefficient pathway flux nodes) is mini- mised by balancing flux throughout the system, and availability of cofactors is balanced. In parallel, the Available online at www.sciencedirect.com ScienceDirect www.sciencedirect.com Current Opinion in Chemical Biology 2017, 40:47–56