The ks domain in an iterative pks can recapture the growing polyketide chain after each round of condensation and terminate polyketide biosynthesis at a precise stage. Here we present the synthesis of unnatural unique polyketide alkaloid hybrid molecules by exploiting the enzyme reaction using precursordirected and structurebased approaches. Growing evidence indicates that the thioesterase te domain is a significant bottleneck within engineered modular pks systems. Polyketide synthases and coenzyme coa ligases were tested to determine the optimal combination of genes for the synthesis of each compound. Polyketide synthase pks enzymes are large multidomain complexes that structurally and. However, very few it1pkss have been discovered in bacteria. Pks polyketide synthase td terminal domain introduction polyketides are a large family of natural products found in bacteria, fungi and plants, and include many clinically important drugs such as tetracycline, daunorubicin, erythromycin, rapamycin and lovastatin. They are biosynthesized from short carboxylic acid precursors by polyketide synthases pkss. Type i polyketide synthases pkss are multifunctional enzymes that are organized into modules, each of which minimally contains a. Unraveling the iterative type i polyketide synthases. The fatty acid synthase fas is a multidomain protein complex maier et al. Structural and evolutionary relationships of atless. Each module incorporates the essential ks, at and acp domains, while all but one include optional reductive activities kr, dh, er.
Engineered biosynthesis of regioselectively modified. Cheng yg, tang gl, shen b 2003 type i polyketide synthase requiring a discrete acyltransferase for polyketide biosynthesis. Users can submit their protein sequences in fasta format to predict whether the given protein sequence belongs to type iii polyketide synthase or not. Here we present the synthesis of unnatural unique polyketidealkaloid hybrid molecules by exploiting the enzyme reaction using precursordirected and structurebased approaches. Polyketide natural products constitute a broad class of compounds with diverse structural features and biological activities. The polyketide synthase gene an15g07920 was known in aspergillus niger cbs 5. The mycolatecontaining compounds constitute major pathogenicity factors of mycobacterium tuberculosis.
An analysis of these genes may provide new targets for genetic improvement to increase relevant secondary metabolites and enhance the plant tolerance to biotic and. Phylogenomic analysis of type i polyketide synthase genes. Bioglass crystallization of type iii polyketide synthase. Kavli institute for theoretical physics, kohn hall, ucsb, santa barbara, ca 93106, usa.
Benzophenone synthase and chalcone synthase from hypercum androsaemum cell cultures. Ossamycin cluster sequence file is available from the. Dalmanol biosyntheses require coupling of two separate. The streptomyces glaucescens tcmkl polyketide synthase. Here, we report the discovery and structure elucidation of the nocap nocardiosisassociated polyketide aglycone by first fully reconstituting the nocap synthase in vitro from purified. Polyketides are alarge family of pharmaceutically important natural products,and the structural modification of. They are produced as secondary metabolites primarily by bacteria and fungi hopwood 1997. Polyketide natural products include many clinically important drugs such as erythromycin antibacterial, epothilone anticancer, rapamycin immunosuppressant, and lovastatin antihypercholesterolemic.
This enzyme catalyzes the condensation of malonyl coa and enzymebound acyl group to afford complex fatty. The biosyntheses of polyketides share striking similarities with fatty acid biosynthesis the pks genes for a certain polyketide are usually organized in one operon or in gene clusters. The polyketide synthase pks catalyzes a novel mechanism. Each type i polyketide synthase module consists of several domains with defined functions, separated by short spacer regions. The biosyntheses of polyketides share striking similarities with fatty acid biosynthesis. Division of pharmaceutical sciences, university of wisconsinmadison, madison, wi 53705, usa. Polyketide synthase an overview sciencedirect topics. The sequencestructurefunction relationship of polyketide biosynthesis. Structural biochemistrypolyketide synthases wikibooks. Surprisingly, the modular polyketide synthase uses only.
Tropane alkaloids biosynthesis involves an unusual type. Several nocardia strains associated with nocardiosis, a potentially lifethreatening disease, house a nonamodular assembly line polyketide synthase pks that presumably synthesizes an unknown polyketide. The order of modules and domains of a complete polyketide synthase is as follows in the order nterminus to cterminus. We were able to produce various phoslactomycin derivatives with altered side. Modular polyketide synthase pks enzymatic complexes contain conserved ketoacyl synthase ks domains, and most pks biosynthetic pathways exceed 30 kb in size. Identification of a putative polyketide synthase gene involved. Here, we report the discovery and structure elucidation of the nocap nocardiosisassociated polyketide aglycone by first fully reconstituting the nocap synthase in vitro from purified protein components followed by heterologous expression in e. Structural and evolutionary relationships of atless type i polyketide synthase ketosynthases.
Complete reconstitution and deorphanization of the 3 mda. Inclusion of c 2acetate in the growth medium resulted in the c enrichment of c8c9, c12c, c14c15, c18c19, c20c21, c26c27, and c32c33, consistent with the incorporation of seven malonylcoa extender units into the rapamycin polyketide core fig. Accordingly, we created a series of hybrid pks modules bearing exchanged te domains from heterologous pathways, and challenged them in vitro with both native and non. The biosynthetic pathway to ossamycin, a macrocyclic polyketide. Chalcone synthase like proteins are comparatively small proteins with a single polypeptide chain and are involved in the biosynthesis of precursors for flavonoids. Engineering a polyketide with a longer chain by insertion of an extra module into the erythromycinproducing polyketide synthase. Polyketide synthase pks the pks family share a number of characteristics with the family of fatty acid synthases fas. Polyketide synthase pathway discovery from soil metagenomic. Debs 6deoxyerythronolide b synthase is a type 1 polyketide synthase responsible for the production of the macrolide 6deoxyerythronolide b 6deb, a aglycone precursor of the antibiotic erythromycin. The origins of specificity in polyketide synthase protein interactions mukund thattai1, yoram burak2, boris i. Mechanistic insights and unsolved problems chaitan khosla, daniel herschlag, david e. Structural and evolutionary relationships of atless type i. Request pdf advances in polyketide synthase structure and function recent progress in the understanding of polyketide synthase pks continues to fuel the growth of combinatorial biosynthesis. Synthesis of three bioactive aromatic compounds by.
Acp is the acyl carrier protein, at acyltransferase, ks ketosynthase, and kr0 a ketoreductase homologue that lacks nadphdependent. Following each condensation reaction, the new betaketo group may be reduced, dehydrated, and reduced again. Engineering modular polyketide synthases for production of. Even once the threedimensional organization of a single polyketide synthase module is resolved, there is still no experimentally based structural prototype for a bimodular not to mention a multimodular polyketide synthase. Phylogenomic analysis of type i polyketide synthase genes in pathogenic and saprobic ascomycetes scott kroken, n. A polyketide synthase encoded by the gene an15g07920 is. Their biosynthetic machinery, represented by type i polyketide synthases pkss, has an architecture in which successive modules catalyse twocarbon linear extensions. High diversity of polyketide synthase genes and the melanin. To isolate a key polyketide biosynthetic intermediate for the 16. They are biosynthesized from acyl coa precursors by polyketide synthases pkss. It is commonly found in the actinomycete bacteria, saccharopolyspora erythraea.
An iterative type i polyketide synthase initiates the. Lohman, tao liu, and ben shen, department of chemistry, the scripps research institute, jupiter, fl 33458, usa. National centre for biological sciences, uas campus, bellary road, bangalore 560065, india. A fosmid metagenomic library constructed from soil at the hancock agricultural research station in hancock, wi, 18,432 clones, average insert of 42 kb was spotted onto a nylon membrane. Type iii polyketide synthases are important for the biosynthesis of flavonoids and various plant polyphenols. Pdf structure of a modular polyketide synthase researchgate.
Structure of a modular polyketide synthase article pdf available in nature 5107506. Unlike all other pkss, these proteins do not have a phosphopantetheinyl ppant arm on which the growing polyketide chains are tethered. Prokaryotic and eukaryotic pkss form a wide variety of biomolecules, including longchain fatty acids, pigments, immunosuppressants, and antibiotics monaghan and tkacz, 1990. We have created phoslactomycin polyketide synthase. Sa response pathways using a fungal polyketide synthase pks gene. Williamsa polyketides are a large group of structurally diverse secondary metabolites with significant therapeutic utility. Genome resequencing analysis revealed that the gene an15g07920 is also present in the ochratoxinproducing a. Rifdh10, the dehydratase domain from the terminal module of the rifamycin polyketide synthase, catalyzes the stereospecific syn dehydration of the model substrate 2s,3s2methyl3hydroxypentanoylrifacp10, resulting in the exclusive formation of e2methyl2pentenoylrifacp10.
We also tested several constructs and identified the best one for increasing levels of endogenous substrates for chorismate, anthranilate, and malonylcoa. Pdf polyketidebased pharmaceuticals are some of our most. Micacocidin is a bacterial natural product with pronounced activity against the human pathogen mycoplasma pneumoniae. Polyketide synthases pkss are one of the most profound. The origins of specificity in polyketide synthase protein. Type i polyketide synthases may have evolved through.
Polyketide synthases pkss are a family of multidomain enzymes or enzyme complexes that produce polyketides, a large class of secondary metabolites, in bacteria, fungi, plants, and a few animal lineages. Advances in polyketide synthase structure and function. Analogous to fatty acid synthases fass, polyketide synthases pkss catalyze the biosynthesis of polyketides through repetitive cc bond. The polyketide synthase reactions are shown on top. Understanding substrate selectivity of phoslactomycin. Fungal polyketide synthase pks 286 is a large protein and consists of a single set of module containing a keto synthase ks, an acyltransferase at, a thioesteraseclaisen cyclase tecyc, and two acyl carrier protein acp domains. Structural basis of a broadly selective acyltransferase.
High diversity of polyketide synthase genes and the melanin biosynthesis gene cluster in penicillium marneffei patrick c. Polyketide synthases pkss are structurally and functionally related to fatty acid synthases fass, as both enzyme classes catalyzes the condensation of activated primary metabolites acetylcoa and malonylcoa to form bketoacetyl polymers linked to the enzyme by thioester bonds. Polyketide synthases produce compounds of high structural and functional diversity. Research article open access unexpected link between polyketide synthase and calcium carbonate biomineralization motoki hojo1,6, ai omi1,7, gen hamanaka2, kazutoshi shindo3, atsuko shimada1, mariko kondo4, takanori narita1,8, masato kiyomoto2, yohei katsuyama5, yasuo ohnishi5, naoki irie1 and hiroyuki takeda1 abstract. Mulberry plants have abundant polyphenols, but very little is known about the mulberry type iii polyketide synthase genes. Shraiman2 1 national centre for biological sciences, bangalore, india, 2 kavli institute for theoretical physics, university of california santa barbara, santa barbara, california, united. Polyketide synthase type iii isolated from uncultured deep. Polyketide synthase secrets revealed june 23, 2014 issue. Jun 20, 20 micacocidin is a bacterial natural product with pronounced activity against the human pathogen mycoplasma pneumoniae. Type iii polyketide synthases in natural product biosynthesis.
Cs bond cleavage by a polyketide synthase domain ming ma, jeremy r. Koch a dissertation submitted in partial fulfillment of the requirements for the degree of doctor of philosophy cancer biology in the university of michigan 2017 doctoral committee. Mutant malonylcoa synthetases with altered specificity for polyketide synthase extender unit generation irina koryakina and gavin j. Engineering polyketide synthase pks pathways that efficiently produce new metabolites requires an understanding of substrate selectivity, flexibility, and sites of failure during chain elongation, processing and termination.
The study is a milestone in the pks field, says polyketide biosynthesis specialist shiouchuan sheryl tsai of the university of california, irvine. Assembly line polyketide synthases acs publications american. The carbon atom source of ta is known from the amino acid ornithine 4,5,6,7 and acetate units 8,9,10 for a long time by elaborate isotopelabeling experiments. The polyketide synthase pks catalyzes a novel mechanism of. Download typeiii polyketide synthase prediction for free. Polyketidepolyketide hybrids are unique natural products with promising bioactivity, but the hybridization processes remain poorly understood. Hspks1 from huperzia serrata is a type iii polyketide synthase pks with remarkable substrate tolerance and catalytic potential. Investigations into the structure and function of type i. Manual inspection and curation using the program artemis 18 led to the defi. Probing selectivity and creating structural diversity through. Mar 14, 2018 structural basis of a broadly selective acyltransferase from the polyketide synthase of splenocin yuan li key laboratory of combinatorial biosynthesis and drug discovery wuhan university, ministry of education, wuhan university school of pharmaceutical sciences, 185 donghu road.
The streptomyces glaucescens tcmkl polyketide synthase and. Despite their diverse end products, pkss are composed of very similar. Polyketides are a large class of structurally and pharmacologically diverse molecules, including many antibiotics and antitumor drugs ohagan 1991. Production of 6methylsalicylic acid by expression of a.
These structures revealed the molecular basis for the stereoselectivity and substrate. Investigations into the structure and function of type i polyketide synthases by aaron a. Pksiiipred is the type iii polyketide synthase prediction server based on svm. In a noniterative modular pks, each module is used only once, and the polyketide intermediate is transferred to a downstream module for chain elongation. The polyketide chains produced by a minimal polyketide synthase are often further derivatized and modified into bioactive natural products palmitic acid is an example of a polyketide since it is formed by the condensation of one acetylcoa primer and seven malonylcoa extender units. Evolutionary and functional analysis of mulberry type iii. Each module within the synthase complex incorporates a single carboxylic acid substrate, derived from a pool of simple precursors, into the pathway product fig. At in the apo form and in complex with benzylmalonyl and pentynylmalonyl extender unit mimics. This is a pdf file of an unedited manuscript that has been. Unexpected link between polyketide synthase and calcium. Department of plant and microbial biology, university of california. Thus, the relative spatial dispositions of adjacent debs modules cannot yet be.
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