Synthetic biology, combinatorial biosynthesis, and chemo‑enzymatic synthesis of isoprenoids

Abstract

Isoprenoids are a large class of natural products with myriad applications as bioactive and commercial compounds. Their diverse structures are derived from the biosynthetic assembly and tailoring of their scaffolds, ultimately constructed from two C5 hemiterpene building blocks. The modular logic of these platforms can be harnessed to improve titers of valuable isoprenoids in diverse hosts and to produce new-to-nature compounds. Often, this process is facilitated by the substrate or product promiscuity of the component enzymes, which can be leveraged to produce novel isoprenoids. To complement rational enhancements and even re-programming of isoprenoid biosynthesis, high-throughput approaches that rely on searching through large enzymatic libraries are being developed. This review summarizes recent advances and strategies related to isoprenoid synthetic biology, combinatorial biosynthesis, and chemo-enzymatic synthesis, focusing on the past 5 years. Emerging applications of cell-free biosynthesis and high-throughput tools are included that culminate in a discussion of the future outlook and perspective of isoprenoid biosynthetic engineering.

Novel Homologs of Isopentenyl Phosphate Kinase Reveal Class‐Wide Substrate Flexibility

“…To begin our investigation, we synthesized a library of 58 alkyl-P analogs (Figure 1A) that included the natural substrate (1), its dimethylallyl isomer (2), and seven previously identified nonnatural substrates of THA/MTH (6-8, 12, 16, 25, 37). [6,[10][11]13] The syntheses were carried out using previously established protocols (see Supplementary Information), [12,14] and the resulting library was divided into four categories based on generalized chemical structures: non-allylic (1,(3)(4)(5)(6)(7)(8)(9)(10)(11), allylic (2, 12-45), benzylic (46-55), and heterocyclic (56)(57)(58). While all groups were designed to probe substrate promiscuity, analogs 2 and 12-58 were synthesized with the utility of their diphosphate products with downstream isoprenoid enzymes (such as PTs) in mind.…”

“…Isoprenoids belong to one the most structurally and chemically diverse classes of natural products in existence and are utilized in a broad range of applications throughout the pharmaceutical and biotechnological industries. [1] Despite their diverse forms and functions, all isoprenoids are derived from the two universal precursors isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), which are derived from either the mevalonate (MVA, Scheme 1A) or deoxy-xylulose-5-phosphate (DXP) pathways in nature. [2] The recent discovery of isopentenyl phosphate kinase (IPK) in Archaea has also led to the identification of an alternate MVA pathway that bifurcates from the classical pathway following the formation of mevalonate-5phosphate (M5P).…”

Advances in Biosynthesis of Natural Products from Marine Microorganisms

“…Likewise, engineered host cells may produce unexpected products or no product at all. Furthermore, the efficiency in reconstructing a biosynthetic system is rather low, especially when substrates with different structural elements are involved [89,114]. While these limitations still exist, combinatorial biosynthesis continues to be a powerful technique in engineering anabolic pathways to produce a wide range of NP derivatives which can be very useful for drug discovery and development.…”

High Versatility of IPP and DMAPP Methyltransferases Enables Synthesis of C6, C7and C8Terpenoid Building Blocks

“…[23][24] For more detailed information about strategies of terpene structure diversification via MTases or other concepts, the reader is referred to recent reviews in this field. [25][26][27] This study aimed at a comparison of the three known IPP MTases and the isolation and characterization of novel C 5 prenyl pyrophosphate MTases with different properties. Detailed descriptions of substrates and products for this group of enzymes and demonstration of their in vivo performance will enable diverse biotechnological applications.…”

“…Due to their ability to extend the chemical space of terpenes, the GPP MTase has been integrated into cellular biosynthesis pathways with different strategies aiming at production of novel terpene products [23–24] . For more detailed information about strategies of terpene structure diversification via MTases or other concepts, the reader is referred to recent reviews in this field [25–27] …”