Enabling Broader Adoption of Biocatalysis in Organic Chemistry.

Biocatalysis is becoming an increasingly impactful method in contemporary synthetic chemistry for target molecule synthesis. The selectivity imparted by enzymes has been leveraged to complete previously intractable chemical transformations and improve synthetic routes toward complex molecules. However, the implementation of biocatalysis in mainstream organic chemistry has been gradual to this point. This is partly due to a set of historical and technological barriers that have prevented chemists from using biocatalysis as a synthetic tool with utility that parallels alternative modes of catalysis. In this Perspective, we discuss these barriers and how they have hindered the adoption of enzyme catalysts into synthetic strategies. We also summarize tools and resources that already enable organic chemists to use biocatalysts. Furthermore, we discuss ways to further lower the barriers for the adoption of biocatalysis by the broader synthetic organic chemistry community through the dissemination of resources, demystifying biocatalytic reactions, and increasing collaboration across the field.

Chemoenzymatic Synthesis of (+)-Xyloketal B.

Xyloketal B is a pentacyclic fungal marine natural product that has shown potential for the treatment of diseases such as Alzheimer's disease and atherosclerosis. Herein, we describe the first asymmetric synthesis of this natural product, which relies on a chemoenzymatic strategy. This approach leverages a biocatalytic benzylic hydroxylation to access to an ortho-quinone methide intermediate which is captured in a [4 + 2] cycloaddition to stereoselectively yield a key cyclic ketal intermediate enroute to (+)-xyloketal B. The relative configuration of this intermediate was rapidly confirmed as the desired stereoisomer using MicroED. To complete the synthesis, a second ortho-quinone methide was accessed through a reductive approach, ultimately leading to the stereoselective synthesis of (+)-xyloketal B.

State-of-the-Art Biocatalysis.

The use of enzyme-mediated reactions has transcended ancient food production to the laboratory synthesis of complex molecules. This evolution has been accelerated by developments in sequencing and DNA synthesis technology, bioinformatic and protein engineering tools, and the increasingly interdisciplinary nature of scientific research. Biocatalysis has become an indispensable tool applied in academic and industrial spheres, enabling synthetic strategies that leverage the exquisite selectivity of enzymes to access target molecules. In this Outlook, we outline the technological advances that have led to the field's current state. Integration of biocatalysis into mainstream synthetic chemistry hinges on increased access to well-characterized enzymes and the permeation of biocatalysis into retrosynthetic logic. Ultimately, we anticipate that biocatalysis is poised to enable the synthesis of increasingly complex molecules at new levels of efficiency and throughput.

Chemoenzymatic Total Synthesis of Natural Products.

The total synthesis of structurally complex natural products has challenged and inspired generations of chemists and remains an exciting area of active research. Despite their history as privileged bioactivity-rich scaffolds, the use of natural products in drug discovery has waned. This shift is driven by their relatively low abundance hindering isolation from natural sources and the challenges presented by their synthesis. Recent developments in biocatalysis have resulted in the application of enzymes for the construction of complex molecules. From the inception of the Narayan lab in 2015, we have focused on harnessing the exquisite selectivity of enzymes alongside contemporary small molecule-based approaches to enable concise chemoenzymatic routes to natural products.We have focused on enzymes from various families that perform selective oxidation reactions. For example, we have targeted xyloketal natural products through a strategy that relies on a chemo- and site-selective biocatalytic hydroxylation. Members of the xyloketal family are characterized by polycyclic ketal cores and demonstrate potent neurological activity. We envisioned assembling a representative xyloketal natural product (xyloketal D) involving a biocatalytically generated ortho-quinone methide intermediate. The non-heme iron (NHI) dependent monooxygenase ClaD was used to perform the benzylic hydroxylation of a resorcinol precursor, the product of which can undergo spontaneous loss of water to form an ortho-quinone methide under mild conditions. This intermediate was trapped using a chiral dienophile to complete the total synthesis of xyloketal D.A second class of biocatalytic oxidation that we have employed in synthesis is the hydroxylative dearomatization of resorcinol compounds using flavin-dependent monooxygenases (FDMOs). We anticipated that the catalyst-controlled site- and stereoselectivity of FDMOs would enable the total synthesis of azaphilone natural products. Azaphilones are bioactive compounds characterized by a pyranoquinone bicyclic core and a fully substituted chiral carbon atom. We leveraged the stereodivergent reactivity of FDMOs AzaH and AfoD to achieve the enantioselective synthesis of trichoflectin enantiomers, deflectin 1a, and lunatoic acid. We also leveraged FDMOs to construct tropolone and sorbicillinoid natural products. Tropolones are a structurally diverse class of bioactive molecules characterized by an aromatic cycloheptatriene core bearing an α-hydroxyketone moiety. We developed a two-step biocatalytic cascade to the tropolone natural product stipitatic aldehyde using the FDMO TropB and a NHI monooxygenase TropC. The FDMO SorbC obtained from the sorbicillin biosynthetic pathway was used in the concise total synthesis of a urea sorbicillinoid natural product.Our long-standing interest in using enzymes to carry out C-H hydroxylation reactions has also been channeled for the late-stage diversification of complex scaffolds. For example, we have used Rieske oxygenases to hydroxylate the tricyclic core common to paralytic shellfish toxins. The systemic toxicity of these compounds can be reduced by adding hydroxyl and sulfate groups, which improves their properties and potential as therapeutic agents. The enzymes SxtT, GxtA, SxtN, and SxtSUL were used to carry out selective C-H hydroxylation and O-sulfation in saxitoxin and related structures. We conclude this Account with a discussion of existing challenges in biocatalysis and ways we can currently address them.

Chemoenzymatic o-Quinone Methide Formation.

Generation of reactive intermediates and interception of these fleeting species under physiological conditions is a common strategy employed by Nature to build molecular complexity. However, selective formation of these species under mild conditions using classical synthetic techniques is an outstanding challenge. Here, we demonstrate the utility of biocatalysis in generating o-quinone methide intermediates with precise chemoselectivity under mild, aqueous conditions. Specifically, α-ketoglutarate-dependent non-heme iron enzymes, CitB and ClaD, are employed to selectively modify benzylic C-H bonds of o-cresol substrates. In this transformation, biocatalytic hydroxylation of a benzylic C-H bond affords a benzylic alcohol product which, under the aqueous reaction conditions, is in equilibrium with the corresponding o-quinone methide. o-Quinone methide interception by a nucleophile or a dienophile allows for one-pot conversion of benzylic C-H bonds into C-C, C-N, C-O, and C-S bonds in chemoenzymatic cascades on preparative scale. The chemoselectivity and mild nature of this platform is showcased here by the selective modification of peptides and chemoenzymatic synthesis of the chroman natural product (-)-xyloketal D.

Au(I)-Catalyzed Synthesis of Trisubstituted Indolizines from 2-Propargyloxypyridines and Methyl Ketones.

A new Au(I)-catalyzed method for the preparation of trisubstituted indolizines from easily accessible 2-propargyloxy-pyridines is reported. The reaction tolerates a wide range of functionality, allowing for diversity to be introduced in four distinct regions of the product (R, R1, R2, and Ar). The proposed mechanism proceeds via enol addition to an allenamide intermediate and explains the observed increase in yields when electron poor methyl ketones are utilized.

Utilizing Native Directing Groups: Mechanistic Understanding of a Direct Arylation Leads to Formation of Tetracyclic Heterocycles via Tandem Intermolecular, Intramolecular C-H Activation.

A mechanistic study on a direct arylation using a native picolylamine directing group is reported. Kinetic studies determined the concentration dependence of substrates and catalysts, as well as catalyst degradation, which led to the development of a new set of reaction conditions capable of affording a robust kinetic profile. During reaction optimization, a small impurity was observed, which was determined to be a dual C-H activation product. A second set of conditions were found to flip the selectivity of the C-H activation to form this tetracycle in high yield. A catalytic cycle is proposed for the intermolecular/intramolecular C-H activation pathway.

Utilizing Native Directing Groups: Synthesis of a Selective IKur Inhibitor, BMS-919373, via a Regioselective C-H Arylation.

BMS-919373 is a highly functionalized quinazoline under investigation as a selective, potent IKur current blocker. By utilizing the aminomethylpyridine side chain at C-4, a selective C-H functionalization at C-5 was invented, enabling the efficient synthesis of this molecule. The strategy of leveraging this inherent directing group allowed the synthesis of this complex heterocycle in only six steps from commodity chemicals. The scope of the C-H activation was further investigated, and the generality of the transformation across a series of bicyclic aromatic heterocycles was explored.

Synthesis of N-Alkenyl 2-Pyridonyl Ethers via a Au(I)-Catalyzed Rearrangement of 2-Propargyloxypyridines.

N-Alkyl 2-pyridones and other enolizable heterocycles are important synthetic constructs, due to their prevalence in natural products and pharmaceutical targets and their capacity to serve as models for a number of biological and chemical processes. The disclosed Au(I)-catalyzed reaction utilizes 2-propargyloxypyridines to access N-alkylated 2-pyridone products derived from both 5-exo and 6-endo addition of the nitrogen to the pendent alkyne. Experimental and computational studies suggest that the desired 5-exo N-alkenyl 2-pyridonyl ethers are formed reversibly in the transformation. After extensive optimization, biaryl Au(I) catalyst 21 was found to overcome the inherent preference for the 6-endo pathway and provide the highest combination of 5-exo selectivity and yield. Herein, we report the application of this new Au(I)-catalyzed C-N bond formation to the preparation of a variety of N-alkenyl 2-pyridonyl ether analogues, which have the potential to serve as an entry point for the synthesis of complex N-alkyl 2-pyridone-containing frameworks.