Biosynthetic Machinery Research Articles

Dynamic imaging of cell wall polysaccharides by metabolic click-mediated labeling of pectins in living elongating cells.

Protein tracking in living plant cells has become routine with the emergence of reporter genes encoding fluorescent tags. Unfortunately, this imaging strategy is not applicable to glycans because they are not directly encoded by the genome. Indeed, complex glycans result from sequential additions and/or removals of monosaccharides by the glycosyltransferases and glycosidases of the cell's biosynthetic machinery. Currently, the imaging of cell wall polymers mainly relies on the use of antibodies or dyes that exhibit variable specificities. However, as immunolocalization typically requires sample fixation, it does not provide access to the dynamics of living cells. The development of click chemistry in plant cell wall biology offers an alternative for live-cell labeling. It consists of the incorporation of a carbohydrate containing a bio-orthogonal chemical reporter into the target polysaccharide using the endogenous biosynthetic machinery of the cell. Once synthesized and deposited in the cell wall, the polysaccharide containing the analog monosaccharide is covalently coupled to an exogenous fluorescent probe. Here, we developed a metabolic click labeling approach which allows the imaging of cell wall polysaccharides in living and elongating cells without affecting cell viability. The protocol was established using the pollen tube, a useful model to follow cell wall dynamics due to its fast and tip-polarized growth, but was also successfully tested on Arabidopsis root cells and root hairs. This method offers the possibility of imaging metabolically incorporated sugars of viable and elongating cells, allowing the study of the long-term dynamics of labeled extracellular polysaccharides.

Biosynthesis of Isonitrile- and Alkyne-Containing Natural Products.

Natural products are a diverse class of biologically produced compounds that participate in fundamental biological processes such as cell signaling, nutrient acquisition, and interference competition. Unique triple-bond functionalities like isonitriles and alkynes often drive bioactivity and may serve as indicators of novel chemical logic and enzymatic machinery. Yet, the biosynthetic underpinnings of these groups remain only partially understood, constraining the opportunity to rationally engineer biomolecules with these functionalities for applications in pharmaceuticals, bioorthogonal chemistry, and other value-added chemical processes. Here, we focus our review on characterized biosynthetic pathways for isonitrile and alkyne functionalities, their bioorthogonal transformations, and prospects for engineering their biosynthetic machinery for biotechnological applications.

An inventory of early branch points in microbial phosphonate biosynthesis.

Microbial phosphonate biosynthetic machinery has been identified in ~5 % of bacterial genomes and encodes natural products like fosfomycin as well as cell surface decorations. Almost all biological phosphonates originate from the rearrangement of phosphoenolpyruvate (PEP) to phosphonopyruvate (PnPy) catalysed by PEP mutase (Ppm), and PnPy is often converted to phosphonoacetaldehyde (PnAA) by PnPy decarboxylase (Ppd). Seven enzymes are known or likely to act on either PnPy or PnAA as early branch points en route to diverse biosynthetic outcomes, and these enzymes may be broadly classified into three reaction types: hydride transfer, aminotransfer, and carbon-carbon bond formation. However, the relative abundance of these branch points in microbial phosphonate biosynthesis is unknown. Also unknown is the proportion of ppm-containing gene neighbourhoods encoding new branch point enzymes and potentially novel phosphonates. In this study we computationally sorted 434 ppm-containing gene neighbourhoods based on these seven branch point enzymes. Unsurprisingly, the majority (56 %) of these pathways encode for production of the common naturally occurring compound 2-aminoethylphosphonate (AEP) or a hydroxylated derivative. The next most abundant genetically encoded intermediates were phosphonoalanine (PnAla, 9.2 %), 2-hydroxyethylphosphonate (HEP, 8.5 %), and phosphonoacetate (PnAc, 6 %). Significantly, about 13 % of the gene neighbourhoods could not be assigned to any of the seven branch points and may encode novel phosphonates. Sequence similarity network analysis revealed families of unusual gene neighbourhoods including possible production of phosphonoacrylate and phosphonofructose, the apparent biosynthetic use of the C-P lyase operon, and a virus-encoded phosphonate. Overall, these results highlight the utility of branch point inventories to identify novel gene neighbourhoods and guide future phosphonate discovery efforts.

Beyond Soil-Dwelling Actinobacteria: Fantastic Antibiotics and Where to Find Them.

Bacterial secondary metabolites represent an invaluable source of bioactive molecules for the pharmaceutical and agrochemical industries. Although screening campaigns for the discovery of new compounds have traditionally been strongly biased towards the study of soil-dwelling Actinobacteria, the current antibiotic resistance and discovery crisis has brought a considerable amount of attention to the study of previously neglected bacterial sources of secondary metabolites. The development and application of new screening, sequencing, genetic manipulation, cultivation and bioinformatic techniques have revealed several other groups of bacteria as producers of striking chemical novelty. Biosynthetic machineries evolved from independent taxonomic origins and under completely different ecological requirements and selective pressures are responsible for these structural innovations. In this review, we summarize the most important discoveries related to secondary metabolites from alternative bacterial sources, trying to provide the reader with a broad perspective on how technical novelties have facilitated the access to the bacterial metabolic dark matter.

Crystal structure and catalytic mechanism of the MbnBC holoenzyme required for methanobactin biosynthesis

Methanobactins (Mbns) are a family of copper-binding peptides involved in copper uptake by methanotrophs, and are potential therapeutic agents for treating diseases characterized by disordered copper accumulation. Mbns are produced via modification of MbnA precursor peptides at cysteine residues catalyzed by the core biosynthetic machinery containing MbnB, an iron-dependent enzyme, and MbnC. However, mechanistic details underlying the catalysis of the MbnBC holoenzyme remain unclear. Here, we present crystal structures of MbnABC complexes from two distinct species, revealing that the leader peptide of the substrate MbnA binds MbnC for recruitment of the MbnBC holoenzyme, while the core peptide of MbnA resides in the catalytic cavity created by the MbnB–MbnC interaction which harbors a unique tri-iron cluster. Ligation of the substrate sulfhydryl group to the tri-iron center achieves a dioxygen-dependent reaction for oxazolone-thioamide installation. Structural analysis of the MbnABC complexes together with functional investigation of MbnB variants identified a conserved catalytic aspartate residue as a general base required for MbnBC-mediated MbnA modification. Together, our study reveals the similar architecture and function of MbnBC complexes from different species, demonstrating an evolutionarily conserved catalytic mechanism of the MbnBC holoenzymes.

Harnessing ortho-Quinone Methides in Natural Product Biosynthesis and Biocatalysis.

The implementation of ortho-quinone methide (o-QM) intermediates in complex molecule assembly represents a remarkably efficient strategy designed by Nature and utilized by synthetic chemists. o-QMs have been taken advantage of in biomimetic syntheses for decades, yet relatively few examples of o-QM-generating enzymes in natural product biosynthetic pathways have been reported. The biosynthetic enzymes that have been discovered thus far exhibit tremendous potential for biocatalytic applications, enabling the selective production of desirable compounds that are otherwise intractable or inherently difficult to achieve by traditional synthetic methods. Characterization of this biosynthetic machinery has the potential to shine a light on new enzymes capable of similar chemistry on diverse substrates, thus expanding our knowledge of Nature's catalytic repertoire. The presently known o-QM-generating enzymes include flavin-dependent oxidases, hetero-Diels-Alderases, S-adenosyl-l-methionine-dependent pericyclases, and α-ketoglutarate-dependent nonheme iron enzymes. In this review, we discuss their diverse enzymatic mechanisms and potential as biocatalysts in constructing natural product molecules such as cannabinoids.

Structural diversity, biosynthesis, and function of plant falcarin-type polyacetylenic lipids.

The polyacetylenic lipids falcarinol, falcarindiol, and associated derivatives, termed falcarins, have a widespread taxonomical distribution in the plant kingdom and have received increasing interest for their demonstrated health-promoting properties as anti-cancer and anti-inflammatory agents. These fatty acid-derived compounds are also linked to plant pathogen resistance through their potent antimicrobial properties. Falcarin-type polyacetylenes, which contain two conjugated triple bonds, are derived from structural modifications of the common fatty acid oleic acid. In the past half century, much progress has been made in understanding the structural diversity of falcarins in the plant kingdom, whereas limited progress has been made on elucidating falcarin function in plant-pathogen interactions. More recently, an understanding of the biosynthetic machinery underlying falcarin biosynthesis has emerged. This review provides a concise summary of the current state of knowledge on falcarin structural diversity, biosynthesis, and plant defense properties. We also present major unanswered questions about falcarin biosynthesis and function.

Ribosome-independent peptide biosynthesis: the challenge of a unifying nomenclature.

The first machineries for non-ribosomal peptide (NRP) biosynthesis were uncovered over 50 years ago, and the dissection of these megasynthetases set the stage for the nomenclature system that has been used ever since. Although the number of exceptions to the canonical biosynthetic pathways has surged in the intervening years, the NRP synthetase (NRPS) classification system has remained relatively unchanged. This has led to the exclusion of many biosynthetic pathways whose biosynthetic machineries violate the classical rules for NRP assembly, and ultimately to a rupture in the field of NRP biosynthesis. In an attempt to unify the classification of NRP pathways and to facilitate the communication within the research field, we propose a revised framework for grouping ribosome-independent peptide biosynthetic pathways based on recognizable commonalities in their biosynthetic logic. Importantly, the framework can be further refined as needed.

Choline transporter-like protein 2 interacts with chitin synthase 1 and is involved in insect cuticle development

Chitin is an aminopolysaccharide present in insects as a major structural component of the cuticle. However, current knowledge on the chitin biosynthetic machinery, especially its constituents and mechanism, is limited. Using three independent binding assays, including co-immunoprecipitation, split-ubiquitin membrane yeast two-hybrid assay, and pull-down assay, we demonstrate that choline transporter-like protein 2 (Ctl2) interacts with krotzkopf verkehrt (kkv) in Drosophila melanogaster. The global knockdown of Ctl2 by RNA interference (RNAi) induced lethality at the larval stage. Tissue-specific RNAi to silence Ctl2 in the tracheal system and in the epidermis of the flies resulted in lethality at the first larval instar. The knockdown of Ctl2 in wings led to shrunken wings containing accumulated fluid. Calcofluor White staining demonstrated reduced chitin content in the first longitudinal vein of Ctl2 knockdown wings. The pro-cuticle, which was thinner compared to wildtype, exhibited a reduced number of chitin laminar layers. Phylogenetic analyses revealed orthologues of Ctl2 in different insect orders with highly conserved domains. Our findings provide new insights into cuticle formation, wherein Ctl2 plays an important role as a chitin-synthase interacting protein.

Engineered biosynthesis of alkyne-tagged polyketides.

Polyketides have demonstrated their significance as therapeutics, industrial products, pesticides, and biological probes following intense study over the past decades. Tagging polyketides with a bioorthogonal functionality enables various applications such as diversification, quantification, visualization and mode-of-action elucidation. The terminal alkyne moiety, as a small, stable and highly selective clickable functionality, is widely adopted in tagging natural products. De novo biosynthesis of alkyne-tagged polyketides offers the unique advantage of reducing the background from feeding the biorthogonal moiety itself, leading to the accomplishment of in situ generation of a clickable functionality for bioorthogonal reactions. Here, we introduce several engineering strategies to apply terminal alkyne biosynthetic machinery, represented by JamABC, which produces a short terminal alkyne-bearing fatty acyl chain on a carrier protein, to functions with different downstream polyketide synthases (PKSs). Successful results in engineering type III and type I PKSs provide engineering guidelines and strategies that are applicable to additional PKSs to produce targeted alkyne-tagged metabolites for chemical and biological applications.

Research highlights on contributions of mitochondrial DNA microsatellite instability in solid cancers - an overview.

Cancer has been broadly considered a genetic disease involving mutations in nuclear DNA and the mitochondrial genome (mtDNA). Mitochondria are essential bioenergetics and biosynthetic machinery found in most eukaryotic organisms. Thus, failure of their function is crucial for tumourigenesis, tumour cell growth, and metastasis. Mitochondrial dysregulation can occur as a consequence of molecular alterations in mtDNA, such as point mutations, deletions, inversions, microsatellite instability (MSI), and copy number variations. This review article aims to highlight the published research work on alterations in mtDNA, with a particular focus on mitochondrial MSI (mtMSI) in various types of solid cancers. Databases including PubMed, Scopus, and Google Scholar were searched for articles about mtMSI and its link to solid cancer published from 1990 till 2021. In this review, we briefly summarize the knowledge related to possible molecular mechanisms causing mtMSI formation and the available information on mtMSI frequency values in all main solid cancer types. Mutations in the mitochondrial genome are widely believed to have a broad impact across various cancers. Based on the available published data, mtMSI can act as a vital risk factor and a potential marker for cancer progression. Further research is required to unravel the role of mtMSI in tumourigenesis.

Dearomative logic in natural product total synthesis.

Covering: 2011 to 2022The natural world is a prolific source of some of the most interesting, rare, and complex molecules known, harnessing sophisticated biosynthetic machinery evolved over billions of years for their production. Many of these natural products represent high-value targets of total synthesis, either for their desirable biological activities or for their beautiful structures outright; yet, the high sp3-character often present in nature's molecules imparts significant topological complexity that pushes the limits of contemporary synthetic technology. Dearomatization is a foundational strategy for generating such intricacy from simple materials that has undergone considerable maturation in recent years. This review highlights the recent achievements in the field of dearomative methodology, with a focus on natural product total synthesis and retrosynthetic analysis. Disconnection guidelines and a three-phase dearomative logic are described, and a spotlight is given to nature's use of dearomatization in the biosynthesis of various classes of natural products. Synthetic studies from 2011 to 2021 are reviewed, and 425 references are cited.

Metabolism and accumulation of hydroxylated fatty acids by castor (Ricinus comunis) seed microsomes

Castor beans accumulate large amounts of triacylglycerols (TAGs) in the seed endosperm. This oil contains hydroxylated ricinoleic levels close to 90%, which is unique among oil seeds. The capacity to accumulate such high levels of such an unusual fatty acids is due to its specific accumulation and channeling. Here, the ability of the castor biosynthetic machinery to accumulate unusual fatty acids in the form of TAGs was investigated, focusing on ricinoleic acid and the structurally analogous lesquerolic and coriolic fatty acids. The metabolism of different radioactive precursors in active membrane fractions from castor bean's were studied, and the rates and accumulation of these fatty acids provided evidence of the different mechanisms involved in the accumulation of hydroxylated fatty acids in this species. In particular, these studies highlighted the potential of castor to accumulate unusual fatty acids other than ricinoleic acid, showing that castor endosperm can efficiently accumulate lesquerolic acid.

Identification and Analysis of the Biosynthetic Gene Cluster for the Indolizidine Alkaloid Iminimycin in Streptomyces griseus.

Indolizidine alkaloids, which have versatile bioactivities, are produced by various organisms. Although the biosynthesis of some indolizidine alkaloids has been studied, the enzymatic machinery for their biosynthesis in Streptomyces remains elusive. Here, we report the identification and analysis of the biosynthetic gene cluster for iminimycin, an indolizidine alkaloid with a 6-5-3 tricyclic system containing an iminium cation from Streptomyces griseus. The gene cluster has 22 genes, including four genes encoding polyketide synthases (PKSs), which consist of eight modules in total. In vitro analysis of the first module revealed that its acyltransferase domain selects malonyl-CoA, although predicted to select methylmalonyl-CoA. Inactivation of seven tailoring enzyme-encoding genes and structural elucidation of four compounds accumulated in mutants provided important insights into iminimycin biosynthesis, although some of these compounds appeared to be shunt products. This study expands our knowledge of the biosynthetic machinery of indolizidine alkaloids and the enzymatic chemistry of PKS.

Oxylipin biosynthesis in spikemoss Selaginella moellendorffii: Identification of allene oxide synthase (CYP74L2) and hydroperoxide lyase (CYP74L1)

Nonclassical P450s of the CYP74 family catalyse the secondary conversions of fatty acid hydroperoxides to bioactive oxylipins in plants. The model organism, spikemoss Selaginella moellendorffii Hieron, possesses at least ten CYP74 genes of novel J, K, L, and M subfamilies. The cloning of three CYP74L genes and catalytic properties of recombinant proteins are described in the present work. The CYP74L1 possessed mainly hydroperoxide lyase (HPL) activity towards the 13(S)-hydroperoxide of α−linolenic acids (13-HPOT) and nearly equal HPL and allene oxide synthase (AOS) activities towards the 13(S)-hydroperoxide of linoleic acids (13-HPOD). The 9-hydroperoxides were poor substrates for CYP74L1 and led to the production of mainly the α−ketols (AOS products) and minorities of HPL and epoxyalcohol synthase (EAS) products. The CYP74L2 possessed the AOS activity towards all tested hydroperoxides. CYP74L3 possessed low HPL/EAS activity. Besides, the aerial parts of S. moellendorffii plants possessed complex oxylipins patterns including divinyl ethers, epoxyalcohols, and 12-oxo-phytodienoic acid. Characterization of the CYP74L enzymes and oxylipin pattern updates the knowledge on the complex oxylipin biosynthetic machinery in the surviving oldest taxa of vascular plants.

Discovery of an Antarctic Ascidian-Associated Uncultivated Verrucomicrobia with Antimelanoma Palmerolide Biosynthetic Potential.

ABSTRACTThe Antarctic marine ecosystem harbors a wealth of biological and chemical innovation that has risen in concert over millennia since the isolation of the continent and formation of the Antarctic circumpolar current. Scientific inquiry into the novelty of marine natural products produced by Antarctic benthic invertebrates led to the discovery of a bioactive macrolide, palmerolide A, that has specific activity against melanoma and holds considerable promise as an anticancer therapeutic. While this compound was isolated from the Antarctic ascidian Synoicum adareanum, its biosynthesis has since been hypothesized to be microbially mediated, given structural similarities to microbially produced hybrid nonribosomal peptide-polyketide macrolides. Here, we describe a metagenome-enabled investigation aimed at identifying the biosynthetic gene cluster (BGC) and palmerolide A-producing organism. A 74-kbp candidate BGC encoding the multimodular enzymatic machinery (hybrid type I-trans-AT polyketide synthase-nonribosomal peptide synthetase and tailoring functional domains) was identified and found to harbor key features predicted as necessary for palmerolide A biosynthesis. Surveys of ascidian microbiome samples targeting the candidate BGC revealed a high correlation between palmerolide gene targets and a single 16S rRNA gene variant (R = 0.83 to 0.99). Through repeated rounds of metagenome sequencing followed by binning contigs into metagenome-assembled genomes, we were able to retrieve a nearly complete genome (10 contigs) of the BGC-producing organism, a novel verrucomicrobium within the Opitutaceae family that we propose here as “Candidatus Synoicihabitans palmerolidicus.” The refined genome assembly harbors five highly similar BGC copies, along with structural and functional features that shed light on the host-associated nature of this unique bacterium.IMPORTANCE Palmerolide A has potential as a chemotherapeutic agent to target melanoma. We interrogated the microbiome of the Antarctic ascidian, Synoicum adareanum, using a cultivation-independent high-throughput sequencing and bioinformatic strategy. The metagenome-encoded biosynthetic machinery predicted to produce palmerolide A was found to be associated with the genome of a member of the S. adareanum core microbiome. Phylogenomic analysis suggests the organism represents a new deeply branching genus, “Candidatus Synoicihabitans palmerolidicus,” in the Opitutaceae family of the Verrucomicrobia phylum. The Ca. Synoicihabitans palmerolidicus 4.29-Mb genome encodes a repertoire of carbohydrate-utilizing and transport pathways, a chemotaxis system, flagellar biosynthetic capacity, and other regulatory elements enabling its ascidian-associated lifestyle. The palmerolide producer’s genome also contains five distinct copies of the large palmerolide biosynthetic gene cluster that may provide structural complexity of palmerolide variants.

A uridine diphosphate-glycosyltransferase GFUGT88A1 derived from edible mushroom Grifola frondosa extends oligosaccharide chains

Fungal glucans play a key role to provide energy, support cell structure, and present biological functions. However, the biosynthetic machineries of fungal glucan chains remain to be elucidated. Hence, we aimed to elucidate the biochemical and catalytic characteristics of a novel glucan synthesis-associated enzyme uridine diphosphate (UDP)-glycosyltransferase GFUGT88A1 derived from Grifola frondosa and predict its potential catalytic mechanism. The purified recombinant GFUGT88A1 had a molecular weight of 51.7 kDa with an optimum temperature of 37 °C and pH of 7.0. GFUGT88A1 showed preference for oligosaccharides with relatively higher polymerization degrees (≥6) as acceptors to extend its chain when using UDP-glucose as a donor. Molecular modeling and docking of GFUGT88A1 with donor UDP-glucose, and acceptors laminaribiose, laminarihexaose, and laminarinonaose indicated that the predicted amino acid residues at the active sites may bind the donor/acceptor protein primarily via hydrogen bonds with various binding energy values. The present study revealed that GFUGT88A1 extended glucan chains, which provides a reference to understand the biosynthesis pathway of mushroom glucans.

Comparative analysis of cuticular waxes of the gametophyte and sporophyte of three species of Polytrichaceae

Few data are available about cuticular wax of mosses, in general, as well as for Polytrichaceae. In addition, most of these studies investigated only the gametophyte. To improve understanding of wax components and their ecological roles, we compared the morphology and the chemical composition of cuticular waxes of the gametophyte and the sporophyte of the Brazilian species Pogonatum pensilvanicum, Polytrichadelphus pseudopolytrichum (endemic), and Polytrichum juniperinum. The waxes were extracted with dichloromethane, derivatized and analyzed using gas chromatography coupled with mass spectrometry. The total wax content was expressed in µg cm–2 and the compounds were identified based on mass fragmentation patterns. The morphology of epicuticular waxes was studied by scanning electron microscopy. Thicker wax layers were observed over the leafy gametophyte and the calyptra, ranging from 0.2 to 0.3 µg cm–2. For the sporophyte, the wax layer was not homogeneous, ranging from 0.01 to 0.04 µg cm–2 over the seta and from 0.07 to 0.2 µg cm–2 over the capsule. While alkanes and fatty acids were found in the two generations, primary alcohols were detected only in the leaves, and aldehydes and esters were found only in the sporophyte. Nonacosan-10-ol was more abundant in the calyptra and capsule. Film waxes were predominant in the gametophyte and sporophyte. Qualitative and quantitative differences were detected in wax composition according to life phases. As far as we know, this is the first time that the sporophyte parts were investigated separately. We hypothesized that the higher wax content in the calyptra may contribute to the protection of the capsule; however, this idea needs to be confirmed. Besides, our data support previous statements that the biosynthetic machinery for wax components was present in the earliest land plants, protecting both the gametophyte and the sporophyte.

Structure and Biosynthesis of Myxofacyclines: Unique Myxobacterial Polyketides Featuring Varing and Rare Heterocycles[] *.

A metabolome‐guided screening approach in the novel myxobacterium Corallococcus sp. MCy9072 resulted in the isolation of the unprecedented natural product myxofacycline A, which features a rare isoxazole substructure. Identification and genomic investigation of additional producers alongside targeted gene inactivation experiments and heterologous expression of the corresponding biosynthetic gene cluster in the host Myxococcus xanthus DK1622 confirmed a noncanonical megaenzyme complex as the biosynthetic origin of myxofacycline A. Induced expression of the respective genes led to significantly increased production titers enabling the identification of six further members of the myxofacycline natural product family. Whereas myxofacyclines A–D display an isoxazole substructure, intriguingly myxofacyclines E and F were found to contain 4‐pyrimidinole, a heterocycle unprecedented in natural products. Lastly, myxofacycline G features another rare 1,2‐dihydropyrol‐3‐one moiety. In addition to a full structure elucidation, we report the underlying biosynthetic machinery and present a rationale for the formation of all myxofacyclines. Unexpectedly, an extraordinary polyketide synthase‐nonribosomal peptide synthetase hybrid was found to produce all three types of heterocycle in these natural products.

Modulation of Specialized Metabolite Production in Genetically Engineered Streptomyces pactum.

Filamentous soil bacteria are known to produce diverse specialized metabolites. Despite having enormous potential as a source of pharmaceuticals, they often produce bioactive metabolites at low titers. Here, we show that inactivation of the pactamycin, NFAT-133, and conglobatin biosynthetic pathways in Streptomyces pactum ATCC 27456 significantly increases the production of the mitochondrial electron transport inhibitors piericidins. Similarly, inactivation of the pactamycin, NFAT-133, and piericidin pathways significantly increases the production of the heat-shock protein (Hsp) 90 inhibitor conglobatin. In addition, four new conglobatin analogues (B2, B3, F1, and F2) with altered polyketide backbones, together with the known analogue conglobatin B1, were identified in this mutant, indicating that the conglobatin biosynthetic machinery is promiscuous toward different substrates. Among the new conglobatin analogues, conglobatin F2 showed enhanced antitumor activity against HeLa and NCI-H460 cancer cell lines compared to conglobatin. Conglobatin F2 also inhibits colony formation of HeLa cells in a dose-dependent manner. Molecular modeling studies suggest that the new conglobatins bind to human Hsp90 and disrupt Hsp90/Cdc37 chaperone/co-chaperone interactions in the same manner as conglobatin. The study also showed that genes that are involved in piericidin biosynthesis are clustered in two different loci located distantly in the S. pactum genome.