Triumphs and Challenges of Natural Product Discovery in the Postgenomic Era.

Natural products have played significant roles as medicine and food throughout human history. Here, we first provide a brief historical overview of natural products, their classification and biosynthetic origins, and the microbiological and genetic methods used for their discovery. We also describe and discuss the technologies that revolutionized the field, which transitioned from classic genetics to genome-centric discovery approximately two decades ago. We then highlight the most recent advancements and approaches in the current postgenomic era, in which genome mining is a standard operation and high-throughput analytical methods allow parallel discovery of genes and molecules at an unprecedented pace. Finally, we discuss the new challenges faced by the field of natural products and the future of systematic heterologous expression and strain-independent discovery, which promises to deliver more molecules in vials than ever before.

Biofuels for a sustainable future.

Rapid increases of energy consumption and human dependency on fossil fuels have led to the accumulation of greenhouse gases and consequently, climate change. As such, major efforts have been taken to develop, test, and adopt clean renewable fuel alternatives. Production of bioethanol and biodiesel from crops is well developed, while other feedstock resources and processes have also shown high potential to provide efficient and cost-effective alternatives, such as landfill and plastic waste conversion, algal photosynthesis, as well as electrochemical carbon fixation. In addition, the downstream microbial fermentation can be further engineered to not only increase the product yield but also expand the chemical space of biofuels through the rational design and fine-tuning of biosynthetic pathways toward the realization of "designer fuels" and diverse future applications.

A deep catalogue of protein-coding variation in 983,578 individuals.

Rare coding variants that substantially affect function provide insights into the biology of a gene1-3. However, ascertaining the frequency of such variants requires large sample sizes4-8. Here we present a catalogue of human protein-coding variation, derived from exome sequencing of 983,578 individuals across diverse populations. In total, 23% of the Regeneron Genetics Center Million Exome (RGC-ME) data come from individuals of African, East Asian, Indigenous American, Middle Eastern and South Asian ancestry. The catalogue includes more than 10.4 million missense and 1.1 million predicted loss-of-function (pLOF) variants. We identify individuals with rare biallelic pLOF variants in 4,848 genes, 1,751 of which have not been previously reported. From precise quantitative estimates of selection against heterozygous loss of function (LOF), we identify 3,988 LOF-intolerant genes, including 86 that were previously assessed as tolerant and 1,153 that lack established disease annotation. We also define regions of missense depletion at high resolution. Notably, 1,482 genes have regions that are depleted of missense variants despite being tolerant of pLOF variants. Finally, we estimate that 3% of individuals have a clinically actionable genetic variant, and that 11,773 variants reported in ClinVar with unknown significance are likely to be deleterious cryptic splice sites. To facilitate variant interpretation and genetics-informed precision medicine, we make this resource of coding variation from the RGC-ME dataset publicly accessible through a variant allele frequency browser.

Complete integration of carbene-transfer chemistry into biosynthesis.

Biosynthesis is an environmentally benign and renewable approach that can be used to produce a broad range of natural and, in some cases, new-to-nature products. However, biology lacks many of the reactions that are available to synthetic chemists, resulting in a narrower scope of accessible products when using biosynthesis rather than synthetic chemistry. A prime example of such chemistry is carbene-transfer reactions1. Although it was recently shown that carbene-transfer reactions can be performed in a cell and used for biosynthesis2,3, carbene donors and unnatural cofactors needed to be added exogenously and transported into cells to effect the desired reactions, precluding cost-effective scale-up of the biosynthesis process with these reactions. Here we report the access to a diazo ester carbene precursor by cellular metabolism and a microbial platform for introducing unnatural carbene-transfer reactions into biosynthesis. The α-diazoester azaserine was produced by expressing a biosynthetic gene cluster in Streptomyces albus. The intracellularly produced azaserine was used as a carbene donor to cyclopropanate another intracellularly produced molecule-styrene. The reaction was catalysed by engineered P450 mutants containing a native cofactor with excellent diastereoselectivity and a moderate yield. Our study establishes a scalable, microbial platform for conducting intracellular abiological carbene-transfer reactions to functionalize a range of natural and new-to-nature products and expands the scope of organic products that can be produced by cellular metabolism.

Genotyping, sequencing and analysis of 140,000 adults from Mexico City.

The Mexico City Prospective Study is a prospective cohort of more than 150,000 adults recruited two decades ago from the urban districts of Coyoacán and Iztapalapa in Mexico City1. Here we generated genotype and exome-sequencing data for all individuals and whole-genome sequencing data for 9,950 selected individuals. We describe high levels of relatedness and substantial heterogeneity in ancestry composition across individuals. Most sequenced individuals had admixed Indigenous American, European and African ancestry, with extensive admixture from Indigenous populations in central, southern and southeastern Mexico. Indigenous Mexican segments of the genome had lower levels of coding variation but an excess of homozygous loss-of-function variants compared with segments of African and European origin. We estimated ancestry-specific allele frequencies at 142 million genomic variants, with an effective sample size of 91,856 for Indigenous Mexican ancestry at exome variants, all available through a public browser. Using whole-genome sequencing, we developed an imputation reference panel that outperforms existing panels at common variants in individuals with high proportions of central, southern and southeastern Indigenous Mexican ancestry. Our work illustrates the value of genetic studies in diverse populations and provides foundational imputation and allele frequency resources for future genetic studies in Mexico and in the United States, where the Hispanic/Latino population is predominantly of Mexican descent.

MIBiG 3.0: a community-driven effort to annotate experimentally validated biosynthetic gene clusters.

With an ever-increasing amount of (meta)genomic data being deposited in sequence databases, (meta)genome mining for natural product biosynthetic pathways occupies a critical role in the discovery of novel pharmaceutical drugs, crop protection agents and biomaterials. The genes that encode these pathways are often organised into biosynthetic gene clusters (BGCs). In 2015, we defined the Minimum Information about a Biosynthetic Gene cluster (MIBiG): a standardised data format that describes the minimally required information to uniquely characterise a BGC. We simultaneously constructed an accompanying online database of BGCs, which has since been widely used by the community as a reference dataset for BGCs and was expanded to 2021 entries in 2019 (MIBiG 2.0). Here, we describe MIBiG 3.0, a database update comprising large-scale validation and re-annotation of existing entries and 661 new entries. Particular attention was paid to the annotation of compound structures and biological activities, as well as protein domain selectivities. Together, these new features keep the database up-to-date, and will provide new opportunities for the scientific community to use its freely available data, e.g. for the training of new machine learning models to predict sequence-structure-function relationships for diverse natural products. MIBiG 3.0 is accessible online at https://mibig.secondarymetabolites.org/.

Enrichment of Immune Dysregulation Disorders in Adult Patients with Human Inborn Errors of Immunity.

Human inborn errors of immunity (IEI) comprise a group of diseases resulting from molecular variants that compromise innate and adaptive immunity. Clinical features of IEI patients are dominated by susceptibility to a spectrum of infectious diseases, as well as autoimmune, autoinflammatory, allergic, and malignant phenotypes that usually appear in childhood, which is when the diagnosis is typically made. However, some IEI patients are identified in adulthood due to symptomatic delay of the disease or other reasons that prevent the request for a molecular study. The application of next-generation sequencing (NGS) as a diagnostic technique has given rise to an ever-increasing identification of IEI-monogenic causes, thus improving the diagnostic yield and facilitating the possibility of personalized treatment. This work was a retrospective study of 173 adults with IEI suspicion that were sequenced between 2005 and 2023. Sanger, targeted gene-panel, and whole exome sequencing were used for molecular diagnosis. Disease-causing variants were identified in 44 of 173 (25.43%) patients. The clinical phenotype of these 44 patients was mostly related to infection susceptibility (63.64%). An enrichment of immune dysregulation diseases was found when cohorts with molecular diagnosis were compared to those without. Immune dysregulation disorders, group 4 from the International Union of Immunological Societies Expert Committee (IUIS), were the most prevalent among these adult patients. Immune dysregulation as a new item in the Jeffrey Model Foundation warning signs for adults significantly increases the sensitivity for the identification of patients with an IEI-producing molecular defect.

Pullenvalenes A-D: Nitric Oxide-Mediated Transcriptional Activation (NOMETA) Enables Discovery of Triterpene Aminoglycosides from Australian Termite Nest-Derived Fungi.

We report on the use of nitric oxide-mediated transcriptional activation (NOMETA) as an innovative means to detect and access new classes of microbial natural products encoded within silent biosynthetic gene clusters. A small library of termite nest- and mangrove-derived fungi and actinomyces was subjected to cultivation profiling using a miniaturized 24-well format approach (MATRIX) in the presence and absence of nitric oxide, with the resulting metabolomes subjected to comparative chemical analysis using UPLC-DAD and GNPS molecular networking. This strategy prompted study of Talaromyces sp. CMB-TN6F and Coccidiodes sp. CMB-TN39F, leading to discovery of the triterpene glycoside pullenvalenes A-D (1-4), featuring an unprecedented triterpene carbon skeleton and rare 6-O-methyl-N-acetyl-d-glucosaminyl glycoside residues. Structure elucidation of 1-4 was achieved by a combination of detailed spectroscopic analysis, chemical degradation, derivatization and synthesis, and biosynthetic considerations.

Novel MECP2 gene therapy is effective in a multicenter study using two mouse models of Rett syndrome and is safe in non-human primates.

The AAV9 gene therapy vector presented in this study is safe in mice and non-human primates and highly efficacious without causing overexpression toxicity, a major challenge for clinical translation of Rett syndrome gene therapy vectors to date. Our team designed a new truncated methyl-CpG-binding protein 2 (MECP2) promoter allowing widespread expression of MECP2 in mice and non-human primates after a single injection into the cerebrospinal fluid without causing overexpression symptoms up to 18 months after injection. Additionally, this new vector is highly efficacious at lower doses compared with previous constructs as demonstrated in extensive efficacy studies performed by two independent laboratories in two different Rett syndrome mouse models carrying either a knockout or one of the most frequent human mutations of Mecp2. Overall, data from this multicenter study highlight the efficacy and safety of this gene therapy construct, making it a promising candidate for first-in-human studies to treat Rett syndrome.

Assessment of a venous thromboembolism prophylaxis shared decision-making intervention (DASH-TOP) using the decisional conflict scale: a mixed-method study.

BACKGROUND: Venous thromboembolism (VTE) in pregnancy is a major cause of maternal morbidity and death. The use of low-molecular-weight heparin (LMWH), despite being the standard of care to prevent VTE, comes with some challenges. Shared decision-making (SDM) interventions are recommended to support patients and clinicians in making preference-sensitive decisions. The quality of the SDM process has been widely assessed with the decisional conflict scale (DCS). Our aim is to report participants' perspectives of each of the components of an SDM intervention (DASH-TOP) in relation to the different subscales of the DCS. METHODS: Design: A convergent, parallel, mixed-methods design. PARTICIPANTS: The sample consisted of 22 health care professionals, students of an Applied Clinical Research in Health Sciences (ICACS) master program. INTERVENTION: We randomly divided the participants in three groups: Group 1 received one component (evidence -based information), Group 2 received two components (first component and value elicitation exercises), and Group 3 received all three components (the first two and a decision analysis recommendation) of the SDM intervention. ANALYSIS: For the quantitative strand, we used a non-parametric test to analyze the differences in the DCS subscales between the three groups. For the qualitative strand, we conducted a content analysis using the decisional conflict domains to deductively categorize the responses. RESULTS: Groups that received more intervention components experienced less conflict and better decision-making quality, although the differences between groups were not statistically significant. The decision analysis recommendation improved the efficacy with the decision-making process, however there are some challenges when implementing it in clinical practice. The uncertainty subscale showed a high decisional conflict for all three groups; contributing factors included low certainty of the evidence-based information provided and a perceived small effect of the drug to reduce the risk of a VTE event. CONCLUSIONS: The DASH-TOP intervention reduced decisional conflict in the decision -making process, with decision analysis being the most effective component to improve the quality of the decision. There is a need for more implementation research to improve the delivery of SDM interventions in the clinical encounter.

Geometric Structures Induced by Deformations of the Legendre Transform.

The recent link discovered between generalized Legendre transforms and non-dually flat statistical manifolds suggests a fundamental reason behind the ubiquity of Rényi's divergence and entropy in a wide range of physical phenomena. However, these early findings still provide little intuition on the nature of this relationship and its implications for physical systems. Here we shed new light on the Legendre transform by revealing the consequences of its deformation via symplectic geometry and complexification. These findings reveal a novel common framework that leads to a principled and unified understanding of physical systems that are not well-described by classic information-theoretic quantities.

A computational framework to explore large-scale biosynthetic diversity.

Genome mining has become a key technology to exploit natural product diversity. Although initially performed on a single-genome basis, the process is now being scaled up to mine entire genera, strain collections and microbiomes. However, no bioinformatic framework is currently available for effectively analyzing datasets of this size and complexity. In the present study, a streamlined computational workflow is provided, consisting of two new software tools: the 'biosynthetic gene similarity clustering and prospecting engine' (BiG-SCAPE), which facilitates fast and interactive sequence similarity network analysis of biosynthetic gene clusters and gene cluster families; and the 'core analysis of syntenic orthologues to prioritize natural product gene clusters' (CORASON), which elucidates phylogenetic relationships within and across these families. BiG-SCAPE is validated by correlating its output to metabolomic data across 363 actinobacterial strains and the discovery potential of CORASON is demonstrated by comprehensively mapping biosynthetic diversity across a range of detoxin/rimosamide-related gene cluster families, culminating in the characterization of seven detoxin analogues.

Skeletal muscle type-specific mitochondrial adaptation to high-fat diet relies on differential autophagy modulation.

In obesity, skeletal muscle mitochondrial activity changes to cope with increased nutrient availability. Autophagy has been proposed as an essential mechanism involved in the regulation of mitochondrial metabolism. Still, the contribution of autophagy to mitochondrial adaptations in skeletal muscle during obesity is unknown. Here, we show that in response to high-fat diet (HFD) feeding, distinct skeletal muscles in mice exhibit differentially regulated autophagy that may modulate mitochondrial activity. We observed that after 4 and 40 weeks of high-fat diet feeding, OXPHOS subunits and mitochondrial DNA content increased in the oxidative soleus muscle. However, in gastrocnemius muscle, which has a mixed fiber-type composition, the mitochondrial mass increased only after 40 weeks of HFD feeding. Interestingly, fatty acid-supported mitochondrial respiration was enhanced in gastrocnemius, but not in soleus muscle after a 4-week HFD feeding. This increased metabolic profile in gastrocnemius was paralleled by preserving autophagy flux, while autophagy flux in soleus was reduced. To determine the role of autophagy in this differential response, we used an autophagy-deficient mouse model with partial deletion of Atg7 specifically in skeletal muscle (SkM-Atg7+/- mice). We observed that Atg7 reduction resulted in diminished autophagic flux in skeletal muscle, alongside blunting the HFD-induced increase in fatty acid-supported mitochondrial respiration observed in gastrocnemius. Remarkably, SkM-Atg7+/- mice did not present increased mitochondria accumulation. Altogether, our results show that HFD triggers specific mitochondrial adaptations in skeletal muscles with different fiber type compositions, and that Atg7-mediated autophagy modulates mitochondrial respiratory capacity but not its content in response to an obesogenic diet.

Nomogram for single-pass automated microkeratome graft preparation for ultrathin Descemet stripping automated endothelial keratoplasty.

PURPOSE: To create a nomogram including the translational speed of the microkeratome blade, microkeratome head size and precut tissue thickness to predict the postcut thickness for Descemet stripping automated endothelial keratoplasty to obtain the thinnest possible graft. METHODS: This prospective study incorporated 48 grafts for DSAEK from March 2017 to June 2020. Corneal tissue for DSAEK was prepared by 3 experienced physicians using the Moria Evolution 3E (Moria Inc, Antony, France) microkeratome with 400, 450 and 500 µm head sizes. Precut central corneal thickness was measured with a DGH 550 handheld pachymeter (Pachette 2), taking an average of 3 readings. The microkeratome head was selected according to precut tissue thickness. The selected microkeratome head size was 150 µm less than the donor cornea thickness. Two translational speeds were used for the microkeratome cuts. One month after surgery, the central lenticular thickness was measured with a Visante® Optical Coherence Tomography caliper (Carl Zeiss Meditec Inc, Germany). A descriptive analysis was performed. RESULTS: Forty-eight donor grafts were prepared. Mean graft thickness was 97.58 ± 29.84 µm (range 39-176 µm). Of the 48 samples, central graft thickness was < 120 µm (81.3%) in 39, < 100 µm (58.3%) in 28 and < 80 µm (37.5%) in 18 at 1-month follow-up. There were no statically significant differences between translational speeds. CONCLUSIONS: A nomogram with an automated microkeratome to obtain thin grafts for DSAEK provided good graft thickness results without donor waste.

Registry Assessment of Peripheral Interventional Devices objective performance goals for superficial femoral and popliteal artery peripheral vascular interventions.

BACKGROUND: The Superficial Femoral Artery-Popliteal EvidencE Development Study Group developed contemporary objective performance goals (OPGs) for peripheral vascular interventions (PVI) for superficial femoral artery (SFA)-popliteal artery disease using the Registry Assessment of Peripheral Interventional Devices. METHODS: The Society for Vascular Surgery Vascular Quality Initiative PVI registry from January 2010 to October 2016 was used to develop OPGs based on SFA-popliteal procedures (n = 21,377) for intermittent claudication and critical limb ischemia (CLI). OPGs included 1-year rates for target lesion revascularization (TLR), major amputation, and 1 and 4-year survival rates. OPGs were calculated for the SFA and popliteal arteries and stratified by four treatments: angioplasty alone (percutaneous transluminal angioplasty [PTA]), self-expanding stenting, atherectomy, and any treatment type. Outcomes were illustrated by unadjusted Kaplan-Meier analyses. RESULTS: Cohorts included PTA (n = 7505), stenting (n = 9217), atherectomy (n = 2510) and any treatment (n = 21,377). The mean age was 69 years, 58% were male, 79% were White, and 52% had CLI. The freedom from TLR OPGs at 1 year in the SFA were 80.3% (PTA), 83.2% (stenting), 83.9% (atherectomy), and 81.9% (any treatments). The freedom from TLR OPGs at 1 year in the popliteal were 81.3% (PTA), 81.3% (stenting), 80.2% (atherectomy), and 81.1% (any treatments). The freedom from major amputation OPGs at 1 year after SFA PVI were 93.4% (PTA), 95.7% (stenting), 95.1% (atherectomy), and 94.8% (any treatments). The freedom from major amputation OPG at 1 year after popliteal PVI were 90.5% (PTA), 93.7% (stenting), 91.8% (atherectomy), and 91.8%, (any treatments). The 4-year survival OPGs after SFA PVI were 76% (PTA), 80% (stenting), 82% (atherectomy), and 79% (any treatments), and for the popliteal artery were 72% (PTA), 77% (stenting), 82% (atherectomy), and 75% (any treatment). On a multivariable analysis, which included patient-level, leg-level, and lesion-level covariates, CLI was the single independent factor associated with increased TLR, amputation, and mortality. CONCLUSIONS: The Superficial Femoral Artery-Popliteal EvidencE Development OPGs define a new, contemporary benchmark for SFA-popliteal interventions using a large subset of real-world evidence to inform more efficient peripheral device clinical trial designs to support regulatory and clinical decision-making. It is appropriate to discuss proposals intended for regulatory approval with the US Food and Drug Administration to refine the OPG to match the specific trial population. The OPGs may be updated using coordinated registry networks to assess long-term real-world device performance.

Impact of the Potential Antitumor Agent 2-(4-Hydroxyphenyl) Amino-1,4-Naphthoquinone (Q7) on Vasomotion Is Mediated by the Vascular Endothelium, But Not Vascular Smooth Muscle Cell Metabolism.

ABSTRACT: Vasomotion is defined as rhythmic oscillations in arterial diameter that regulate the blood flow and blood pressure. Because antitumor treatment may impair vascular functions and increase the blood pressure, we sought to evaluate whether a new naphthoquinone derivative, postulated as an antitumor agent, manifests adverse effects on vascular function. In this article, we evaluated the toxicity of 2-(4-hydroxyphenyl) amino-1,4-naphthoquinone (Q7) and its effects on vascular vasomotion in 3 models of vascular structure: endothelial cells, aortic ring, and smooth muscle cells. Although showing nontoxic effects, Q7 inhibited the formation of capillary-like structures of the EA.hy926 endothelial cell line grown on Matrigel. In exvivo experiments with aortic rings precontracted with phenylephrine (PE, 10-6 M), Q7 (10-5 M) significantly (P < 0.05) reduced vascular rhythmic contractions induced by the acetylcholine (ACh; 10-7-10-5 M), whereas sodium nitroprusside (a nitric oxide donor; 10-8 M) recovered the vasomotion. Furthermore, Q7 (10-5 M) did not decrease KCl-induced vascular rhythmic contractions in the aortic rings precontracted with BaCl2 (a nonselective K+ channel blocker; 10-3 M). Vascular smooth muscle cells (A7r5) preincubated with Q7 (10-5 M) for 3 hours also demonstrated a reduced glucose uptake. However, the Adenosine Triphosphate content was unaffected, suggesting that the rapid reduction in vasomotion observed in vascular reactivity experiments did not involve cellular metabolism but may be due to faster mechanisms involving endothelial nitric oxide and K+ channels leading to oscillations in intracellular Ca2+. In summary, the naphthoquinone derivative Q7 presents low cytotoxicity yet may alter the endothelial cell response and vasomotion in the absence of changes in smooth muscle cell metabolism.

Design and health: An interdisciplinary baseline experience of product design for health needs of the Chilean elderly.

Objective: Describing the results of a User-Centered Design workshop using Challenge Based Learning or CBL, where students from Chilean industrial design and health degrees, developed solutions to resolve health problems among the elderly. METHODS: It was a pre-experimental study; 45 university students took part in a workshop: 39 from Industrial Design, 3 from Speech Therapy, 2 from Medical Technology and 1 from Medicine. In this workshop, the students, using CBL in disciplinary heterogeneous groups, faced a 3-week challenge to develop products to overcome a health problem for an elderly person. Once the product was presented, the professors and students assessed the conceptual proposals, using a semantic differential. The students also evaluated the workshop with a perception survey. RESULTS: Both the students and professors positively evaluated the usefulness and functionality of the conceptual proposals; although, the industrial design students were more critical about these aspects. The originality of proposals evaluated was the worst. Regarding the workshop, all students (100%) felt that instructions were clear as were the three moments of design, with problem definition stage best evaluated. Conclusion: The CBL is shown to be an educational tool that allows training professionals in product design and in developing health technology that is suitable for the users' needs. Regarding the process, heterogeneous make-up of the groups and clear external guide appear as essential for CBL to work well.

Structural Mechanism of Regioselectivity in an Unusual Bacterial Acyl-CoA Dehydrogenase.

Terminal alkenes are easily derivatized, making them desirable functional group targets for polyketide synthase (PKS) engineering. However, they are rarely encountered in natural PKS systems. One mechanism for terminal alkene formation in PKSs is through the activity of an acyl-CoA dehydrogenase (ACAD). Herein, we use biochemical and structural analysis to understand the mechanism of terminal alkene formation catalyzed by an γ,δ-ACAD from the biosynthesis of the polyketide natural product FK506, TcsD. While TcsD is homologous to canonical α,ß-ACADs, it acts regioselectively at the γ,δ-position and only on α,ß-unsaturated substrates. Furthermore, this regioselectivity is controlled by a combination of bulky residues in the active site and a lateral shift in the positioning of the FAD cofactor within the enzyme. Substrate modeling suggests that TcsD utilizes a novel set of hydrogen bond donors for substrate activation and positioning, preventing dehydrogenation at the α,ß position of substrates. From the structural and biochemical characterization of TcsD, key residues that contribute to regioselectivity and are unique to the protein family were determined and used to identify other putative γ,δ-ACADs that belong to diverse natural product biosynthetic gene clusters. These predictions are supported by the demonstration that a phylogenetically distant homologue of TcsD also regioselectively oxidizes α,ß-unsaturated substrates. This work exemplifies a powerful approach to understand unique enzymatic reactions and will facilitate future enzyme discovery, inform enzyme engineering, and aid natural product characterization efforts.

Chemoinformatic-Guided Engineering of Polyketide Synthases.

Polyketide synthase (PKS) engineering is an attractive method to generate new molecules such as commodity, fine and specialty chemicals. A significant challenge is re-engineering a partially reductive PKS module to produce a saturated ß-carbon through a reductive loop (RL) exchange. In this work, we sought to establish that chemoinformatics, a field traditionally used in drug discovery, offers a viable strategy for RL exchanges. We first introduced a set of donor RLs of diverse genetic origin and chemical substrates  into the first extension module of the lipomycin PKS (LipPKS1). Product titers of these engineered unimodular PKSs correlated with chemical structure similarity between the substrate of the donor RLs and recipient LipPKS1, reaching a titer of 165 mg/L of short-chain fatty acids produced by the host Streptomyces albus J1074. Expanding this method to larger intermediates that require bimodular communication, we introduced RLs of divergent chemosimilarity into LipPKS2 and determined triketide lactone production. Collectively, we observed a statistically significant correlation between atom pair chemosimilarity and production, establishing a new chemoinformatic method that may aid in the engineering of PKSs to produce desired, unnatural products.