Mechanistic Analysis of a Type II Polyketide Synthase. Role of Conserved Residues in the β-Ketoacyl Synthase−Chain Length Factor Heterodimer

Dreier, Juerg; Khosla, Chaitan

doi:10.1021/bi992121l

Cited by 68 publications

(62 citation statements)

References 22 publications

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“…2). We computationally tested the long-standing hypothesis that the volume of the KS-CLF amphipathic cavity, which houses the growing polyketide chain during biosynthesis, is a main determinant of polyketide chain length (5,(8)(9)(10)(11)(12)(13)(14)(15). Using the solved actinorhodin KS-CLF structure as a template for in silico mutagenesis, we calculated the predicted volume of the KS-CLF cavity for five PKS clades ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Besides the ACP, other genes involved in backbone biosynthesis include acyltransferase (AT) and priming KS (KSIII). Most characterized type II systems are primed with acetyl units and "borrow" these enzymes from the FAS pathway (8)(9)(10)(11)(12)(13)(14)(15)(21)(22)(23). Systems using nonacetate starting units rely on secondary AT and KSIII enzymes (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The remarkable chemical diversity observed in this class of molecules is thought to originate from variations in chain length and tailoring reactions. Previous phylogenetic studies have revealed the role of the CLF in controlling the chain length of type II polyketides (8)(9)(10)(11)(12)(13)(14)(15), but the evolution of the KS-CLF within the context of the entire protein assembly is not well understood.…”

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confidence: 99%

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Evolution of chemical diversity by coordinated gene swaps in type II polyketide gene clusters

Hillenmeyer

Vandova

Berlew

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Natural product biosynthetic pathways generate molecules of enormous structural complexity and exquisitely tuned biological activities. Studies of natural products have led to the discovery of many pharmaceutical agents, particularly antibiotics. Attempts to harness the catalytic prowess of biosynthetic enzyme systems, for both compound discovery and engineering, have been limited by a poor understanding of the evolution of the underlying gene clusters. We developed an approach to study the evolution of biosynthetic genes on a cluster-wide scale, integrating pairwise gene coevolution information with large-scale phylogenetic analysis. We used this method to infer the evolution of type II polyketide gene clusters, tracing the path of evolution from the single ancestor to those gene clusters surviving today. We identified 10 key gene types in these clusters, most of which were swapped in from existing cellular processes and subsequently specialized. The ancestral type II polyketide gene cluster likely comprised a core set of five genes, a roster that expanded and contracted throughout evolution. A key C24 ancestor diversified into major classes of longer and shorter chain length systems, from which a C20 ancestor gave rise to the majority of characterized type II polyketide antibiotics. Our findings reveal that (i) type II polyketide structure is predictable from its gene roster, (ii) only certain gene combinations are compatible, and (iii) gene swaps were likely a key to evolution of chemical diversity. The lessons learned about how natural selection drives polyketide chemical innovation can be applied to the rational design and guided discovery of chemicals with desired structures and properties.evolution | polyketide | natural products | gene cluster

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

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Evolution of chemical diversity by coordinated gene swaps in type II polyketide gene clusters

Hillenmeyer

Vandova

Berlew

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…thesis (33). Therefore the mupirocin biosynthesis machine may employ multiple ACPs to enhance the rate and overcome the rate-limiting step in the most economical way.…”

Section: Role Of Tandemly Repeated Acyl Carrier Proteinsmentioning

confidence: 99%

Tandemly Duplicated Acyl Carrier Proteins, Which Increase Polyketide Antibiotic Production, Can Apparently Function Either in Parallel or in Series

Rahman

Hothersall

Crosby

et al. 2005

Journal of Biological Chemistry

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Polyketide biosynthesis involves the addition of subunits commonly derived from malonate or methylmalonate to a starter unit such as acetate. Type I polyketide synthases are multifunctional polypeptides that contain one or more modules, each of which normally contains all the enzymatic domains for a single round of extension and modification of the polyketide backbone. Acyl carrier proteins (ACP(s)) hold the extender unit to which the starter or growing chain is added. Normally there is one ACP for each ketosynthase module. However, there are an increasing number of known examples of tandemly repeated ACP domains, whose function is as yet unknown. For the doublet and triplet ACP domains in the biosynthetic pathway for the antibiotic mupirocin from Pseudomonas fluorescens NCIMB10586 we have inactivated ACP domains by inframe deletion and amino acid substitution of the active site serine. By deletion analysis each individual ACP from a cluster can provide a basic but reduced activity for the pathway. In the doublet cluster, substitution analysis indicates that the pathway may follow two parallel routes, one via each of the ACPs, thus increasing overall pathway flow. In the triplet cluster, substitution in ACP5 blocked the pathway. Thus ACP5 appears to be arranged "in series" to ACP6 and ACP7. Thus although both the doublet and triplet clusters increase antibiotic production, the mechanisms by which they do this appear to be different and depend specifically on the biosynthetic stage involved. The function of some ACPs may be determined by their location in the protein rather than absolute enzymic activity.

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“…Such enzymes can serve as a mechanistic model for the type II polyketide cyclases that catalyze a series of intramolecular aldol reactions upon enzyme-bound poly-␤-ketone intermediates (27,28). Importantly, this structure shows strong topological similarity to the actva-orf6 monoxygenase in the actinorhodin biosynthetic pathway, which suggests that this fold is ideally suited to serve as a framework for rearrangements and chemical modification of polyaromatic substrates.…”

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confidence: 99%

Structural and Functional Analysis of Tetracenomycin F2 Cyclase from Streptomyces glaucescens

Thompson

Katayama

Watanabe

et al. 2004

Journal of Biological Chemistry

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Tetracenomycin F2 cyclase (tcmI gene product), catalyzes an aromatic rearrangement in the biosynthetic pathway for tetracenomycin C in Streptomyces glaucescens. The x-ray structure of this small enzyme has been determined to 1.9-Å resolution together with an analysis of site-directed mutants of potential catalytic residues. The protein exhibits a dimeric ␤␣␤ ferredoxin-like fold that utilizes strand swapping between subunits in its assembly. The fold is dominated by four strands of antiparallel sheet and a layer of ␣-helices, which creates a cavity that is proposed to be the active site. This type of secondary structural arrangement has been previously observed in polyketide monooxygenases and suggests an evolutionary relationship between enzymes that catalyze adjacent steps in these biosynthetic pathways. Mutational analysis of all of the obvious catalytic bases within the active site suggests that the enzyme functions to steer the chemical outcome of the cyclization rather than providing a specific catalytic group. Together, the structure and functional analysis provide insight into the structural framework necessary to perform the complex rearrangements catalyzed by this class of polyketide cyclases.

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Mechanistic Analysis of a Type II Polyketide Synthase. Role of Conserved Residues in the β-Ketoacyl Synthase−Chain Length Factor Heterodimer

Cited by 68 publications

References 22 publications

Evolution of chemical diversity by coordinated gene swaps in type II polyketide gene clusters

Evolution of chemical diversity by coordinated gene swaps in type II polyketide gene clusters

Tandemly Duplicated Acyl Carrier Proteins, Which Increase Polyketide Antibiotic Production, Can Apparently Function Either in Parallel or in Series

Structural and Functional Analysis of Tetracenomycin F2 Cyclase from Streptomyces glaucescens

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