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Polyketide synthase

Polyketide synthases (PKSs) are a family of multifunctional enzymes belonging to the fatty acid synthase (FAS) superfamily that catalyze the biosynthesis of polyketides, a structurally diverse group of secondary metabolites produced primarily by bacteria, fungi, and plants.^[1] These enzymes assemble polyketide chains through iterative decarboxylative Claisen condensations of acyl units, typically derived from malonyl-CoA or methylmalonyl-CoA, followed by optional reductions, dehydrations, and cyclizations to generate bioactive compounds such as antibiotics (e.g., erythromycin), immunosuppressants, and anticancer agents.^[1] Unlike FAS, which produce uniform fatty acids, PKSs exhibit programmable specificity, allowing for the creation of thousands of distinct polyketide scaffolds with pharmaceutical relevance.^[2] Structurally, PKSs are large multienzyme complexes ranging from 1 to 10 megadaltons, organized as modular assembly lines where functional domains are fused into polypeptides.^[1] Key domains include the ketosynthase (KS) for chain elongation, acyltransferase (AT) for extender unit loading, and acyl carrier protein (ACP) that tethers the growing polyketide chain via a flexible 4'-phosphopantetheine arm.^[2] PKSs are classified into types based on architecture: Type I (modular, multidomain proteins like the 6-deoxyerythronolide B synthase, DEBS), Type II (dissociated enzymes for aromatic polyketides), and Type III (standalone chalcone synthase-like enzymes in plants).^[1] Within Type I, cis-AT PKSs integrate AT domains within each module, while trans-AT PKSs use separate, diffusible AT enzymes, enabling greater structural diversity.^[2] Recent structural studies reveal dimeric, sheet-like organizations stabilized by linker interactions, facilitating efficient chain translocation via mechanisms like the "turnstile" model.^[2]^[3] The evolutionary origins of PKSs trace back to FAS-like ancestors, with diversification driven by gene duplication and domain shuffling, resulting in over 8,000 genomically encoded "orphan" PKS gene clusters awaiting functional characterization.^[1] This vast biosynthetic potential underscores PKSs' role in microbial chemical ecology and their exploitation in biotechnology for engineering novel therapeutics through combinatorial biosynthesis.^[1]

Overview

Definition and function

Polyketide synthases (PKSs) are a family of multi-enzyme complexes that catalyze the biosynthesis of polyketides, a structurally diverse class of secondary metabolites including macrolides such as erythromycin, polyenes such as amphotericin B, and aromatic compounds such as tetracyclines.^[4] These enzymes assemble polyketide backbones through the sequential incorporation of acyl units, resulting in natural products with a wide range of biological activities, including antibiotic, anticancer, and immunosuppressive properties.^[5] The primary function of PKSs centers on iterative decarboxylative Claisen condensation reactions, where acyl-CoA thioesters—most commonly malonyl-CoA—serve as building blocks to form linear β-ketoacyl chains bound to an acyl carrier protein (ACP).^[6] Unlike fully reducing systems, PKSs incorporate optional β-carbon processing steps, such as ketoreduction, dehydration, and enoyl reduction, which can be skipped to retain keto, hydroxyl, or alkene functionalities and generate structural diversity.^[5] The core chain elongation reaction is depicted as:

\text{R-C(O)-S-ACP} + \text{HOOC-CH}_2\text{-C(O)-S-CoA} \rightarrow \text{R-C(O)-CH}_2\text{-C(O)-S-ACP} + \text{CO}_2 + \text{CoA-SH}

This condensation, facilitated by the ketosynthase (KS) domain, decarboxylates the malonyl extender unit to drive carbon-carbon bond formation.^[7] PKSs operate through mechanisms highly analogous to those of fatty acid synthases (FAS), sharing evolutionary origins, precursor pools like malonyl-CoA, and domain architectures for condensation and processing, but PKSs diverge by allowing programmable incomplete reduction of β-keto intermediates to preserve reactive groups essential for polyketide folding and cyclization.^[6] This flexibility enables the production of unsaturated or oxygenated scaffolds, contrasting with the saturated fatty acids generated by FAS.^[8] These enzymes are predominantly found in bacteria, particularly actinomycetes such as Streptomyces species, as well as in fungi and plants; while PKS-like enzymes occur in animals, often as FAS variants producing simpler lipids, recent findings (as of 2024) reveal complex polyketide production in certain invertebrates like molluscs via widespread FAS-like PKS, though such occurrences remain less common than in microbes.^[9]^[10]

Diversity and occurrence

Polyketide synthases (PKSs) produce a vast array of structurally diverse natural products, ranging from linear chains to complex cyclic and aromatic scaffolds. This diversity arises from variations in chain length, degree of reduction, cyclization patterns, and post-assembly modifications, enabling the biosynthesis of macrolactones such as erythromycin, enediyne antibiotics like calicheamicin, and polyaromatic compounds including tetracycline.^[8]^[11]^[12] PKSs are widely distributed across prokaryotes and eukaryotes, with bacterial sources predominating in the production of bioactive metabolites. In bacteria, particularly actinomycetes like Streptomyces species, PKSs synthesize numerous antibiotics and antitumor agents.^[13] Fungal PKSs, such as those in Aspergillus species, generate cholesterol-lowering compounds like lovastatin, and often produce hybrid molecules combining polyketide and non-ribosomal peptide moieties through NRPS-PKS fusion enzymes.^[14]^[15] In plants, type III PKSs, such as chalcone synthase, contribute to phenolic compounds like flavonoids.^[16] Metazoans, including marine sponges, host PKSs primarily through bacterial symbionts that assemble specialized polyketides for chemical defense.^[17]^[18] PKS genes are typically organized into biosynthetic gene clusters or operons, which facilitate coordinated expression and regulation of the multi-enzyme assembly process. These clusters often include accessory genes for tailoring enzymes, transporters, and regulators, ensuring efficient production of the polyketide scaffold.^[19]^[20] The evolutionary diversification of PKSs has been driven by horizontal gene transfer, allowing dissemination across microbial taxa and contributing to their broad phylogenetic distribution. Over 10,000 distinct polyketides have been identified to date, with many more biosynthetic potentials remaining untapped in underexplored genomes.^[21]^[22]^[23]

Classification

Type I PKS

Type I polyketide synthases (PKSs) are large, multifunctional enzymes organized as megasynthases, consisting of covalently linked polypeptides that assemble complex polyketide structures through sequential condensation and modification steps.^[24] These systems are characterized by their modular architecture, where each elongation cycle is mediated by a dedicated module, distinguishing them from the dissociable enzymes of other PKS types. Type I PKSs can operate in either non-iterative (modular) or iterative modes, with the former using each module once for linear chain extension and the latter reusing modules for repeated cycles.^[25] Type I PKSs are subdivided into cis-acyltransferase (cis-AT) and trans-acyltransferase (trans-AT) subtypes based on the organization of the acyltransferase (AT) domain responsible for loading extender units such as malonyl-CoA or methylmalonyl-CoA. In cis-AT systems, the AT domain is integrated within each module and is typically specific for particular substrates, a configuration common in bacteria like actinomycetes.^[24] In contrast, trans-AT systems feature a discrete, standalone AT protein that delivers extender units to multiple modules, providing greater substrate flexibility; these are prevalent in myxobacteria and contribute to the diversity of polyketides produced by these organisms.^[25] Both subtypes share a core module structure comprising ketosynthase (KS), AT, and acyl carrier protein (ACP) domains, with optional reductive domains including ketoreductase (KR), dehydratase (DH), and enoylreductase (ER) that control the oxidation state of the growing chain.^[24] A prototypical example of a cis-AT Type I PKS is the 6-deoxyerythronolide B synthase (DEBS) from Saccharopolyspora erythraea, which assembles the 14-membered macrolactone precursor to the antibiotic erythromycin. DEBS consists of three large polypeptides (DEBS1, DEBS2, DEBS3) encoding six modules in total, initiating with a propionyl-CoA starter unit and extending via six methylmalonyl-CoA units, with specific modules incorporating KR, DH, and ER for partial reduction to achieve the desired stereochemistry and functionality.^[25] Type I PKSs are further classified by reduction level into fully reducing (FR), partially reducing (PR), and non-reducing (NR) subtypes, where FR and PR systems include active reductive domains to produce aliphatic polyketides, while NR systems lack these domains and instead form aromatic compounds through cyclization.^[24] For instance, the iterative NR-PKS 6-methylsalicylic acid synthase (6-MSAS) in fungi like Penicillium patulum synthesizes the simple aromatic polyketide 6-methylsalicylic acid via multiple iterations of malonyl-CoA extension without reduction.^[26]

Type II PKS

Type II polyketide synthases (PKSs) are discrete, multidomain enzymes that function iteratively to biosynthesize aromatic polyketides, primarily in bacteria such as Streptomyces species.^[27] These systems rely on a minimal set of enzymes, including the ketosynthase α (KSα) for chain elongation, the chain length factor (CLF, also known as KSβ) that controls chain length and facilitates aromatization, and the acyl carrier protein (ACP) that tethers the growing polyketide chain.^[28] Accessory enzymes, such as ketoreductases (KR) for β-keto group reduction and aromatase/cyclases (ARO/CYC) for ring folding and aromatization, further modify the intermediate to yield the final aromatic scaffold.^[29] Unlike the large, modular assemblies of Type I PKSs, Type II systems use these dissociated components iteratively for efficient production of structurally diverse aromatics.^[1] The biosynthetic mechanism of Type II PKSs begins with the loading of a starter unit, typically acetyl-CoA, onto the ACP, followed by iterative decarboxylative Claisen condensations with malonyl-CoA units to extend the chain and form a linear poly-β-ketone intermediate.^[30] The KSα-CLF heterodimer catalyzes each elongation step, with the CLF ensuring precise chain length by positioning the ACP-bound substrate.^[31] Post-elongation, the poly-β-keto chain undergoes programmed folding into specific conformations (e.g., C9-C14 or C7-C12), driven by intrinsic reactivity or accessory enzymes like KR and CYC, culminating in dehydration, oxidation, and aromatization to produce fused ring systems such as decalin or anthraquinone cores.^[27] A classic example is the biosynthesis of actinorhodin, a blue-pigmented benzoisochromanequinone antibiotic produced by Streptomyces coelicolor via its minimal PKS (actI-orf1 to orf3), which generates a C16 polyketide chain that cyclizes to the aromatic product.^[32] Similarly, anthracyclines like doxorubicin, an anticancer agent from Streptomyces peucetius, are assembled by a Type II PKS involving iterative condensation to a 21-carbon decaketide, followed by cyclization and post-PKS modifications to form the linear tetracyclic aglycone.^[33] These pathways exemplify how Type II PKSs generate bioactive molecules with applications in pigmentation, antibiotic resistance, and chemotherapy. Type II PKS gene clusters are frequently co-localized with genes encoding tailoring enzymes, such as glycosyltransferases, oxygenases, and methyltransferases, which diversify the polyketide scaffold through post-assembly modifications like glycosylation or halogenation.^[27] A 2025 review underscores the health impacts of Type II-derived compounds, particularly anticancer anthraquinones like those in doxorubicin, which intercalate DNA and inhibit topoisomerase II, though challenges remain in mitigating cardiotoxicity.^[27] Variations in Type II PKS architecture include hybrid systems in certain bacteria, such as those combining Type II and Type III elements to produce extended polyketide chains beyond typical aromatic limits, enabling the synthesis of linear or polyene structures.

Type III PKS

Type III polyketide synthases (PKSs) represent the simplest class within the PKS family, characterized as compact homodimeric enzymes belonging to the chalcone synthase (CHS)-fold superfamily. Unlike types I and II, they lack an acyl carrier protein (ACP) and instead directly utilize free malonyl-CoA as the extender unit, paired with starter substrates such as acetyl-CoA or p-coumaroyl-CoA, to build polyketide chains through iterative decarboxylative condensations. These enzymes are predominantly found in plants and fungi, where they synthesize phenolic compounds, but they also occur in bacteria, contributing to the structural diversity of secondary metabolites.^[34]^[35] The biosynthetic mechanism of type III PKSs occurs within a single active site cavity, employing a conserved cysteine-histidine-asparagine (Cys-His-Asn) catalytic triad to facilitate repeated rounds of condensation, β-keto processing, and cyclization. Typically, these enzymes perform three to four iterative condensations, yielding tetraketide products such as chalcones, which undergo spontaneous or enzyme-assisted aromatization to form phenolics; longer chains can be produced depending on active site architecture. A prominent example is chalcone synthase (CHS) in plants, which condenses one p-coumaroyl-CoA starter with three malonyl-CoA units to produce naringenin chalcone, the foundational scaffold for flavonoid biosynthesis essential in plant defense and pigmentation. In bacteria, enzymes like 1,3,6,8-tetrahydroxynaphthalene synthase (THNS), such as the RppA homolog in Streptomyces species, catalyze the condensation of four malonyl-CoA units to generate 1,3,6,8-tetrahydroxynaphthalene, a precursor to melanin pigments.^[36] Recent studies highlight the functional flexibility of fungal type III PKSs, which produce diverse metabolites including resorcinols through substrate promiscuity and minimal gene clusters often co-localized with tailoring enzymes like cytochrome P450s. These enzymes exhibit variations in operational modes, primarily decarboxylative condensations leading to aromatic products via aldol or Claisen cyclizations for resorcinol or phloroglucinol scaffolds, respectively, though some retain carboxyl groups in intermediates for alternative chain extensions. For instance, fungal enzymes like AiizPKS demonstrate broad acceptance of fatty acyl-CoA starters (C9-C10), yielding alkylresorcinols with pharmaceutical potential.^[37]^[37]

Structure and components

Modules

In modular polyketide synthases (PKSs), primarily those of Type I, modules serve as the fundamental organizational units that orchestrate the iterative assembly of polyketide chains. Each module is a self-contained functional unit composed of 3 to 7 catalytic domains covalently linked within one or more polypeptides, enabling a processive, assembly-line mechanism where the growing polyketide intermediate remains tethered to the enzyme complex without dissociation between elongation cycles. This modularity ensures efficient substrate channeling and structural diversity in the final products, as the sequence and domain composition of modules dictate the chain length, branching, and modifications.^[38] The overall organization of a modular PKS typically commences with a loading module, which includes an acyltransferase (AT) domain to select and activate a starter unit (such as a propionyl group) and load it onto an acyl carrier protein (ACP) domain, often preceded by a ketosynthase (KS) domain in some systems. This is followed by a series of elongation modules, each minimally containing a KS domain for decarboxylative Claisen condensation, an AT domain to incorporate extender units like malonyl-CoA, and an ACP domain to tether the β-ketoacyl intermediate. Additional optional domains within elongation modules—such as ketoreductase (KR), dehydratase (DH), and enoylreductase (ER)—perform stereospecific reductions and dehydrations on the β-keto group during each cycle. Intermodular transfer of the elongated chain relies on docking domains located at the C-terminus of the upstream ACP and N-terminus of the downstream KS, which mediate specific, non-covalent recognition to ensure ordered progression along the assembly line.^[38]00156-X) A representative example is the pikromycin PKS from Streptomyces venezuelae, which features one loading module and six elongation modules distributed across four polypeptides (PikAI–PikAIV), directing the biosynthesis of polyketide intermediates that cyclize to form 12- or 14-membered macrolactone rings in products like methymycin and pikromycin, respectively. The precise order of these modules determines the carbon chain length and positions of functional groups, highlighting how modularity underpins product specificity. In trans-AT PKS systems, where AT activity is provided by discrete, standalone enzymes rather than integrated domains, specialized intermodular linkers—such as the LINKS motif—facilitate the structural organization of modules into a cohesive megacomplex, and certain variants enable module skipping through mechanisms like alternative translation starts, allowing control over chain extension and final product size.^[39]^[40]^[41]

Domains

Polyketide synthases (PKSs) are composed of multiple catalytic and carrier domains that work in concert to assemble polyketide chains. The core domains include the ketosynthase (KS), acyltransferase (AT) or malonyl/acetyl transferase (MT), and acyl carrier protein (ACP). The KS domain catalyzes the decarboxylative Claisen condensation to form carbon-carbon bonds during chain elongation, utilizing a conserved cysteine residue in its active site to accept the acyl chain from the ACP.^[1] The AT (or MT in fungal systems) domain selects and transfers extender units, such as malonyl-CoA or methylmalonyl-CoA, to the ACP via a serine nucleophile in a two-step transacylation mechanism, determining substrate specificity through sequence motifs in its active site.^[1] The ACP domain serves as the central scaffold, tethering the growing polyketide chain via a 4'-phosphopantetheine (Ppant) arm attached to a serine residue, which enables flexible shuttling of intermediates between other domains over distances up to approximately 18 Å.^[1] Reductive domains modify the β-keto group introduced during each elongation cycle and are optional depending on the desired polyketide structure. The ketoreductase (KR) domain reduces the β-ketoacyl-ACP to a β-hydroxyacyl-ACP using NADPH, exerting stereocontrol over both α- and β-carbon centers, as seen in its ability to epimerize α-methyl groups in certain modules.^[1] The dehydratase (DH) domain subsequently eliminates water from the β-hydroxyacyl-ACP to form a trans- or cis-α,β-unsaturated thioester, featuring a double-hotdog fold that accommodates the substrate.^[1] The enoylreductase (ER) domain then reduces the α,β-enoyl-ACP to the saturated acyl-ACP using NADPH, influencing the stereochemistry of adjacent methyl groups.^[1] Additional domains handle chain release and post-assembly modifications. The thioesterase (TE) domain cleaves the completed polyketide from the ACP through hydrolysis or intramolecular nucleophilic attack, often forming macrocycles, and adopts a dimeric α/β hydrolase fold with a substrate-binding channel.^[1] In non-reducing PKSs (NR-PKSs), cyclase (CYC) and aromatase (ARO) domains promote regiospecific cyclization and aromatization of the polyketide chain, with di-domain ARO/CYC structures revealing N-terminal recognition motifs for chain length specificity.^[42] Domain interactions are mediated by dynamic docking and thioester linkages, exemplified by the ACP's Ppant arm forming thioesters such as ACP-S-C(O)-R with the polyketide intermediate R, which transfers to the KS active site cysteine for condensation. The ACP docks specifically with the KS via its helix II region, positioning the Ppant arm for efficient substrate delivery, as visualized in high-resolution structures. Recent cryo-EM studies (2024) of full-length Type I PKS modules, such as DEBS module 1 and PikAIII, reveal asymmetric megasynthase conformations with two reaction chambers and turnstile-like ACP movements, enabling ordered domain access during biosynthesis.^[43] In engineering contexts, domain swapping—such as exchanging AT or KR domains between modules—has been exploited to alter substrate specificity and stereochemistry, producing hybrid polyketides while preserving overall assembly-line fidelity.^[44] These domains integrate into larger modules to facilitate iterative chain extension, as detailed in the modules section.

Biosynthetic mechanism

Chain initiation and elongation

Chain initiation in polyketide synthases (PKSs) occurs through the loading module, where an acyltransferase (AT) domain selectively transfers a starter unit, such as propionyl-CoA, onto the acyl carrier protein (ACP) via transesterification to form a thioester bond with the ACP's phosphopantetheine arm.^[7] This step primes the growing polyketide chain, with the ACP tethering the intermediate throughout biosynthesis to facilitate subsequent reactions.^[8] In parallel, extender units like malonyl-CoA are loaded onto a separate ACP domain by another AT, setting the stage for chain extension; decarboxylation of this malonyl unit occurs during the subsequent condensation step rather than independently.^[5] Elongation proceeds via the ketosynthase (KS) domain, which catalyzes a decarboxylative Claisen condensation between the ACP-tethered growing chain (initially the starter unit) and the malonyl-ACP extender.^[5] The mechanism involves nucleophilic attack by the enolate derived from the decarboxylated malonyl unit on the carbonyl of the acyl chain, extending the polyketide by two carbons and yielding a β-ketoacyl-ACP intermediate.^[8] This reaction can be represented as:

\text{R-C(O)-S-KS} + \ce{^{-}OOC-CH2-C(O)-S-ACP} \rightarrow \text{R-C(O)-CH2-C(O)-S-ACP} + \ce{CO2}

where R represents the growing polyketide chain transferred to the KS active site cysteine prior to condensation.^[5] The KS ensures stereospecificity in the condensation geometry, typically favoring the formation of the thermodynamically stable β-keto product without introducing new chiral centers at this stage.^[8] These initiation and elongation steps repeat iteratively, with the chain remaining covalently bound to the ACP for 4–20 cycles depending on the PKS system, adding successive two-carbon units to build the polyketide backbone.^[8] In Type I PKSs, each cycle occurs within dedicated modules, with inter-module transfer of the ACP-bound intermediate facilitated by docking domains that ensure processive assembly.^[5] By contrast, Type II and Type III PKSs employ iterative reuse of minimal domain sets (KS, AT, ACP in Type II; standalone active sites in Type III) for multiple rounds of elongation without modular segmentation.^[8]

Modification and termination

In polyketide biosynthesis, post-condensation modifications diversify the growing chain by altering functional groups introduced during elongation. Ketoreductase (KR) domains stereospecifically reduce β-ketoacyl intermediates to β-hydroxyacyl groups, while dehydratase (DH) domains eliminate water to form α,β-unsaturated enoyl groups, and enoylreductase (ER) domains further saturate these to alkyl chains, with the presence or absence of these optional domains determining the oxidation state of the final product. Methyltransferase (MT) domains incorporate methyl groups using S-adenosylmethionine (SAM), often at C- or O-positions to enhance structural complexity, as seen in certain aromatic polyketides. Cyclization mechanisms finalize the polyketide core by forming rings through intramolecular reactions. In non-reducing polyketide synthases (NR-PKS), product template (PT) domains guide regioselective first-ring cyclization via aldol or Claisen condensations, where the regioselectivity—such as C2-C7 versus C2-C11—determines the scaffold, exemplified by C2-C7 aldol cyclization yielding resorcylic acid macrolactones like radicicol.^[45]^[46] Dieckmann condensations, facilitated by reductase-like domains in some NR-PKS, enable macrolide formation by intramolecular attack on the thioester, producing tetramate or lactone rings in compounds like equisetin.^[45] Aromatization follows in many cases, driven by aromatase/cyclase (ARO/CYC) domains in type II PKS, which dehydrate and oxidize polyene intermediates to form phenolic rings, as in the C7-C12 cyclization of actinorhodin.^[27] Termination releases the mature polyketide from the synthase. Thioesterase (TE) domains typically hydrolyze the thioester bond for linear products or catalyze macrolactonization by nucleophilic attack of a hydroxyl on the thioester carbonyl, as in the pikromycin pathway where TE promotes 14-membered ring closure. In hybrid PKS-NRPS systems, TE off-loads the polyketide chain to a non-ribosomal peptide synthetase (NRPS) module for further peptide extension and cyclization.^[45] Type III PKS exhibit auto-termination after 3-4 elongation cycles via spontaneous thioester hydrolysis or intrinsic cyclization, releasing triketide or tetraketide products like acylphloroglucinols without dedicated TE domains.^[47] Recent studies on thioesterase evolution highlight opportunities for engineering selectivity. Directed evolution of a modular PKS TE from the pikromycin pathway identified mutations enhancing macrolactonization of hybrid amide-containing intermediates, achieving a six-fold yield increase for unnatural macrocyclic ring systems and revealing key residues influencing substrate specificity.^[48]

Biological roles

Pharmacological relevance

Polyketides biosynthesized by polyketide synthases (PKS) represent a cornerstone of modern pharmacology, with numerous compounds serving as the basis for FDA-approved therapeutics across multiple disease areas. These natural products and their derivatives exhibit diverse mechanisms of action, targeting bacterial protein synthesis, fungal membranes, DNA replication, and cellular signaling pathways, among others. Their structural complexity and potent bioactivity have made them indispensable in treating infections, cancers, and metabolic disorders, though production challenges persist.^[49] In the realm of antibiotics, PKS-derived polyketides have been pivotal in combating bacterial infections. Macrolides such as erythromycin, produced by type I PKS in Saccharopolyspora erythraea, bind to the 50S subunit of the bacterial ribosome, inhibiting translocation during protein synthesis and exhibiting activity against Gram-positive bacteria.^[49] Tetracyclines, synthesized via type II PKS in Streptomyces species, reversibly bind to the 30S ribosomal subunit, blocking tRNA attachment and halting protein synthesis in a broad spectrum of bacteria, including those causing acne and respiratory infections.^[49] However, rising antibiotic resistance has prompted semi-synthetic modifications; for instance, clarithromycin, derived from erythromycin through methylation, enhances acid stability and tissue penetration while retaining the ribosomal binding mechanism.^[50] Anticancer applications of PKS polyketides leverage their ability to disrupt DNA and cellular architecture. Anthracyclines like doxorubicin, generated by type II PKS in Streptomyces peucetius, intercalate into DNA and inhibit topoisomerase II, leading to DNA damage and apoptosis in rapidly dividing cancer cells; it is widely used in treating leukemias, lymphomas, and solid tumors.^[49] Enediynes, such as calicheamicin produced by Micromonospora echinospora, feature a potent DNA-cleaving warhead activated by thiol reduction, and in clinical use, calicheamicin γ1 is conjugated to antibodies (e.g., in gemtuzumab ozogamicin) for targeted delivery to CD33-positive acute myeloid leukemia cells.^[50] Beyond antimicrobials and cytotoxics, PKS polyketides address immunosuppression and metabolic conditions. Sirolimus (rapamycin), a type I PKS product from Streptomyces hygroscopicus, forms a complex with FKBP12 that allosterically inhibits mTOR, suppressing T-cell proliferation and cytokine production; it is approved for preventing organ transplant rejection and treating lymphangioleiomyomatosis.^[51] Statins like lovastatin, biosynthesized by type I PKS in Aspergillus terreus, competitively inhibit HMG-CoA reductase, the rate-limiting enzyme in cholesterol biosynthesis, reducing cardiovascular risk through lowered LDL levels.^[52] Epothilones, such as ixabepilone (a semi-synthetic derivative of epothilone B from Sorangium cellulosum type I PKS), stabilize microtubules by binding β-tubulin, preventing depolymerization and inducing mitotic arrest; it is indicated for metastatic breast cancer resistant to taxanes.^[53] Over 20 FDA-approved drugs derive from PKS polyketides, underscoring their clinical impact, yet challenges in commercialization remain. Native microbial producers often yield low titers due to complex regulatory networks and slow growth, necessitating fermentation optimization or heterologous expression.^[54] Total chemical synthesis is rare owing to the intricate stereochemistry and macrocyclic architectures, favoring semi-synthesis or biosynthetic engineering for analogs.^[55]

Ecological significance

Polyketides serve critical functions in microbial defense within natural ecosystems, particularly by enabling bacteria to outcompete rivals. Soil-dwelling streptomycetes, such as Streptomyces species, produce polyketide-based antibiotics like tetracycline and erythromycin to inhibit the growth of neighboring bacteria and fungi, thereby securing nutrient resources in nutrient-limited environments.^[56] Similarly, the polyketide antifungal amphotericin B, synthesized by Streptomyces nodosus, targets ergosterol in fungal membranes, providing a competitive edge against fungal antagonists in soil microbiomes.^[57] In addition to defense, polyketides facilitate intercellular signaling and predator deterrence. The blue pigment actinorhodin, produced by Streptomyces coelicolor, acts as a quorum-sensing signal that coordinates antibiotic production and community behaviors in response to environmental cues like iron scarcity, enhancing group-level survival in soil biofilms.^[58] Fungal polyketides such as patulin, emitted by Penicillium expansum, function as toxins that deter predators and suppress competing microbes, promoting fungal dominance in fruit and soil niches.^[59] Polyketides also underpin symbiotic interactions across ecosystems. In lichens, fungal mycobionts produce polyketides like depsides and depsidones via type I PKS, which protect the algal photobiont from biotic stresses and stabilize the mutualistic partnership.^[60] Marine sponge-associated bacteria synthesize polyketides through dedicated PKS pathways, contributing to host defense against pathogens and facilitating nutrient exchange in the symbiosis.^[61] In plants, type III PKS-derived flavonoids, such as those in floral nectaries, attract pollinators by providing visual and olfactory cues, supporting reproductive success and biodiversity.^[62] A 2023 study revealed that fungal secondary metabolism, including polyketide production, is regulated by an RNA-binding protein complex (CsdA/RsdA), which fine-tunes gene expression to adapt to ecological pressures like nutrient availability.^[63] Many polyketide biosynthetic gene clusters remain silent under standard conditions but activate in response to environmental stresses, such as oxidative damage or nutrient limitation, allowing microbes to produce defensive compounds on demand.^[64] Overall, polyketides enhance soil microbiome diversity by modulating microbial interactions and competition, while undiscovered pathways in marine actinomycetes suggest untapped ecological roles in oceanic carbon cycling and symbioses.^[56]^[65]

Engineering and future directions

Synthetic biology approaches

Synthetic biology approaches to polyketide synthase (PKS) engineering leverage the modular architecture of these enzymes to generate novel polyketides with altered structures and properties. By manipulating specific domains or entire modules, researchers can redirect substrate incorporation and chain modification, enabling the production of compounds not found in nature. These strategies often involve genetic recombination techniques to create hybrid enzymes, which are then expressed in suitable host organisms for scalable biosynthesis. Domain and module engineering represent foundational methods for tailoring PKS activity. Swapping acyltransferase (AT) domains alters substrate specificity, allowing incorporation of non-native extender units such as methylmalonyl-CoA instead of malonyl-CoA, as demonstrated in engineered DEBS modules that produced branched-chain polyketides. Similarly, deletion or inactivation of reductive domains, such as ketoreductases (KR) and dehydratases (DH), prevents β-carbon reduction, yielding unreduced polyketide analogs with preserved keto groups, exemplified by modifications to the avermectin PKS that generated fully unsaturated products. Heterologous expression systems facilitate the testing and optimization of engineered PKS constructs outside their native hosts. Escherichia coli and Saccharomyces cerevisiae are commonly used due to their genetic tractability and established metabolic engineering tools, though challenges like improper protein folding and assembly of large multidomain PKSs persist in these prokaryotic and eukaryotic systems, respectively. A prominent example is the expression of the 6-deoxyerythronolide B synthase (DEBS) in engineered E. coli strains, which produces the erythromycin precursor 6-dEB at titers exceeding 1 g/L after pathway optimization, including co-expression of propionyl-CoA carboxylase for precursor supply. Combinatorial biosynthesis expands structural diversity by mixing modules from disparate PKS pathways, creating hybrid synthases capable of novel chain elongations. For instance, swapping modules between the erythromycin (DEBS) and tylosin PKSs has yielded macrolides with modified structures, enhancing potency against resistant bacterial strains. This approach exploits inter-module interfaces to maintain catalytic fidelity, though compatibility issues can reduce yields. Recent innovations include gene conversion techniques for iterative PKS editing, inspired by natural evolutionary mechanisms, enabling successive domain replacements without disrupting overall synthase integrity. A 2025 study demonstrated this method's application to modular PKSs, achieving up to threefold improvements in analog production through targeted homology-directed repairs. Additionally, fusions between non-ribosomal peptide synthetases (NRPS) and PKS modules generate hybrid polyketide-peptide molecules, as seen in engineered NRPS-PKS chimeras that incorporate amino acids into polyketide backbones for enhanced bioactivity. CRISPR-based tools have accelerated PKS engineering by enabling precise cluster activation and modification. CRISPR-Cas9 knock-in strategies activate silent biosynthetic gene clusters (BGCs) in streptomycetes by inserting strong promoters upstream of PKS genes, boosting titers of macrolides like erythromycin by up to 6-fold.^[66] These methods address regulatory barriers but require careful design to avoid off-target effects in complex actinomycete genomes.

Recent advances

Recent advances in structural biology have significantly enhanced understanding of polyketide synthase (PKS) architecture, particularly through the integration of cryo-electron microscopy (cryo-EM) and AlphaFold predictions for full-length Type I modular PKS models. In 2024, high-resolution cryo-EM structures of enzymes like Lsd14 and DEBS module 1 revealed asymmetric domain arrangements and flexible linkers that facilitate substrate channeling and conformational dynamics during biosynthesis.^[43] Similarly, AlphaFold-guided mutagenesis confirmed docking mechanisms between acyl carrier protein (ACP) domains and downstream ketosynthases, highlighting the role of flexible inter-domain linkers in modular assembly-line efficiency.^[67] Computational tools have accelerated PKS engineering by enabling retrobiosynthetic design of chimeric systems. The BioPKS pipeline, introduced in 2025, automates the integration of multifunctional Type I PKS modules with monofunctional enzymes to predict and construct novel polyketide pathways, demonstrating successful in silico targeting of complex carbon backbones.^[68] Complementing this, directed evolution strategies targeting thioesterase (TE) domains have improved product yields in modular PKS by optimizing release mechanisms, as shown in high-throughput assessments of pentaketide synthases.^[40] New metabolite discoveries underscore the biosynthetic versatility of PKS enzymes. In 2025, studies revealed functional flexibility in Type III PKS, enabling iterative elongation to produce diverse structures beyond canonical flavonoids, including those with ecological roles in microbial interactions.^[69] Concurrently, genomic analysis linked a PKS gene cluster (pks1) in Trichoderma atroviride to the biosynthesis of 6-pentyl-α-pyrone, a bioactive compound with antifungal properties, confirming its essential role via gene knockout experiments.^[70] Engineering breakthroughs have expanded PKS applications toward sustainable chemicals. In 2024, repurposing a fully reducing PKS module from a bacterial system achieved de novo production of 2-methyl Guerbet-like lipids, precursors for biofuels, with titers exceeding 100 mg/L in engineered Escherichia coli.^[71] Regulatory insights from fungal systems further advanced control mechanisms; a 2023 study identified an RNA-binding protein complex (CsdA/RsdA) that governs secondary metabolism by modulating PKS gene expression in Aspergillus fumigatus, offering targets for enhanced metabolite yields.^[72] A 2025 review on Type II PKS emphasized their health impacts, detailing how aromatic polyketides like anthracyclines contribute to anticancer therapies while addressing gaps in engineering novel substrates from marine-derived clusters for broader therapeutic applications.^[27]

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